CA1124644A - Organo-polymeric particles forming lattice by organic polymer adhesive - Google Patents

Abstract

An element for the analysis or transport of liquid, especially aqueous liquids, contains a struc-ture comprising a plurality of heat-stable, organo-polymeric particles nonswellable in and impermeable to the liquid, and an adhesive concentrated at particle surface areas contiguous adjacent particles, bonding the particles into a coherent, three-dimensional lat-tice which is nonswellable in the liquid. A substan-tial portion of the particle surface area in this lat-tice structure is therefore effectively free from adhesive. The lattice structure has interconnected void spaces among the particles representing a total void volume of about 25 to 80 percent to provide for transport of the liquid. The adhesive comprises an organic polymer different from that of the particles and insoluble in the liquid under analysis. The amount of adhesive in the structure is less than 10 weight percent of the particles.
The particulate structure of these elements can contain interactive compositions useful for the analysis of various substances in liquids, especially high-molecular-weight proteinaceous substances in aqueous biological liquids. Multizone elements con-taining, in fluid contact, at least two zones having a particulate structure as described above or one such zone together with other functional zones are also disclosed.

Description

ORGANO-POLYMERIC PARTIC~S FORMING LATTICE
BY ORGANIC POLYMER ADHESIVE
F~eld of the Invention The present invention relates to elements having a particulate structure effec~ive for transport or analysis of liquids. These structures are particularly useful in '1dry chemistry" analysis of aqueous liquids. "Dry chemistry"
analysis refers to analytical methods and techniques which are carried out using chemical reagents contained in various "dry-to-the-touch" test elements such as "dip-and-read" test strips, mutilayer test elements and the like.
Background of the Inven~ion An increasingly large number of analytical tests, procedures and analyses (i.e., assays) must be performed each day on many kinds of liquid samples including, but not limited to, aqueous biological ~luids such as blood, serum, urine, cerebrospinal fluid and the like. To handle and meet critical laboratory needs effectively, "dry chemistry" ana-lytical elements used in these analyses should function rap-idly, require minimal operator involvement, provide accurate and reproducible results, and reduce the severe fluid-handling problems presented by the very nature of liquid samples.
Prior to the present invention, certain improved "dry chemistry" multilayer analytical elements were devel~
oped, as described in US Patent 3,992,158, to overcome many o~ the foregoing problems.
Nevertheless, large complex molecules and cellular structures contained in many aqueous liquid samples or used as reagents (hereinafter termed "interactive compositions") in many liquid analysis procedures create particular diEfi-culty in the design and development of "dry chemistry" ana-lytical elements. These substances tend to clog and impede Eluid flow in conventional liquid-transport structures con-tained in many analytical elements.
The present invention provides a novel particulate structure for the transport or analysis oE liquids which readily accommodates and transports many lar~e, complex molecules and cells which may be contained in such liquids.

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Related Art The "dry chemistry" elements Or U.S. Patent 3~992,158 provide a highly eff~ective multilayer element ror analysis of liquids, especlally aqueous biological liquids.
5 These elements have an isotropically porous, non-fibrous spreading layer and a reagent layer. The spreading layer acts as an aqueous liquid transport layer.
The spreading layers of U.S. Patent 3,992,158 can be prepared from a variety o~ materials including a "blushed"
10 polymer material or a particulate material. In the case of a particulate material, spreading layer porosity is created by interconnected open (i.e., void) spaces among the parti-cles. Pigments, diatomaceous earth particles~ microcrystal-line colloid materials, and spherical particles of uniform 1~ size, such as resinous or glass beads, represent useful particulate materials ~or such spreadin~ layers. Two specific types of particulate structures disclosed ln the patent are structures composed of self-adhesive particulate materials and structures containing particulate materials 20 and a separate binder as an adhesive.
In the case o~ particulate materials which can be rendered self-adhesive, such as heat-softenable polymer particles, one can form a porous structure as follows: A
plurality of the particles are heat- or solvent-sof~ened and

2~ compacted to form a layer of agglomerated particles in which ad~acent particles are fused together at points of lnter-particle contact. U.S. Patent 2,297,248 issued September 29, 1942 and U.S. Patent 2,745,141 issued May 15, 1956 disclose speci~ic particulate structures prepared in this manner.

3 U.S. Patent 2,297,248 discloses porous filter elements prepared by compacting a plurality o~ acrylic polymer particles under heat or solvent action and pressure so that adJacent particles of the ~ilter element are fused to one another at points of interparticle contact. U.S. Patent 35 2,745,141 discloses a particulate structure made by spraying thermoplastic particles~ e.g., polyethylene or polystyrene particles, through a heat zone onto a constant speed movlng base. The exterior sur~ace only o~ the particles becomes molten and fuses ad~acent particles together ~d to the base.
~ .S. Patent 3,5743150 issued April 6, 1971, represents another variant Or a par~iculate structure 5 compos-ed of selr-adhesive particles. This patent describes an open pore polyurethane structure composed o~ coherent spherical polyurethane particles of less than 10 mlcrons ln diameter. The structure is formed in sltu on a sultable support by precipitating the spherical polyurethane partlcles l-~from a dilute mixture o~ polyurethane-forming reactants dispersed in an organic diluent that serves as a non-solvent for the particulate polyurethane reaction product.
A second specific type o~ particulate structure described ln U.S. Patent 3,992,158 is that composed o~
15 particles bonded together by a separate adhesive. For example, this patent describes a porous structure composed o~ non-adherent particles, such as glass beads, coated wlth a thin adherent layer of a hydrophillc colloid, e.g., gelatin or poly(vinyl alcohol). When the collold coating 2~` dries, the resultant porous layer structure formed by the adjacent particles retains its integrity and maintalns sufficient open spaces among component particles to permit passage of aqueous liquids. U.S. Patent 2,297,248 discloses that filter elements~ also, can be prepared by adhering 2~ together adjacent particles of~a particulate structure with a "suitable cement." However, the patent provides no examples of such filter elements and no description or examples of "suitable cements."
The general class o~ non-ribrous spreadlng layer 3 structures described in U.S. Patent 3,99~,158 can ef`fectlvely transport aqueous liquids as well as a variety of substances contained in such liquids. Nevertheless~ improvements in the specific particulate layer structures Or the types discussed above would be hlghly desirable to provide struc-3~tures which are capable o~ transportlng large, complex molecules, for example, macromolecules of biological origin, and cells, for example, red blood cells, that are contained in body fluids.

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In structures composed Or self-adherent particles as described ln U.S. Patent 3,992,158 and the other patents noted above, the heat- or solvent-so~tened particles tend to readily a~glomerate and fill in the interparticle open spaces o~ the structure. Thus, high molecular weight substances employed in many aqueous liquid assays readily clog and impede rluid rlow in such structures.
Similarly, many structures composed of particles bonded together w~th a separate cement or blnder in the manner broadly disclosed in U.S. Patents 3,992,158 and 2,297,248 tend to become clogged and impede ~luid flow Or liquids containing complex, high molecular weight substances.
For example, in the course o~ work relatlng to the present invention, many structures composed of particles and a separate adhesive were found to exhiblt the problem o~
having open spaces clogged and filled in. One apparent cause of this problem is that a substantial layer of adhesive distributed over most or all of the particle surface area in the structure can lead to "open space fill-in" by the adhesive. In addition, because of their solubility 9 many common adhesives, e.g., water-soluble colloids and other water-soluble polymers such as poly(vinyl pyrolidone), exhibit reduced adhesive strength in the presence of aqueous liquids. Also9 many particulate materials, e.g. 9 cellulosic 25 particles, tend to swell in the presence of aqueous liquids.
Accordingly, when a particulate structure prepared from these materials is used to analyze aqueous liquids, struc ture coherency is reduced or lost~ and partial or complete "open space fill-in" occurs.
30 Summary of the Invention The present invention provides an element having an improved particulate structure for the analysis or trans-port of liquid. This structure can readily accommodate many high molecular weight substances, lncluding red blood cells, 35 dissolved or dispersed in liquid samples or interactive compositions used in liquld analysis procedures without clogging or otherwise substantially impeding ~luid transport in the element. Accordingly J the elements of the invention represent highly effective transport structures for liquids .
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containing complex, high molecular weight substances o~
analytical interest. In particular, these elements repre-sent highly effective structures ~or use ln aqueous liquid analyses which require fluid migration of complex, high molecular wel~ht substances within the ele~ent during the analysis procedure.
The elements of the invention can perform a highly e~ficient "spreading" function ~or liquids containing either low or high molecular weight substances of analytlcal 10 interest, hereinafter termed analytes, especially high molecular weight analytes. That is, these elements have a particulate structure which can readily take up, uni~ormly distribute within itself, meter, and rapidly transport applied liquid samples containing any of a wide variety of 15 analytes In these respects, the elements of the inventlon perform the same highly useful "spreading" function provided by conventional non-fibrous, particulate spreading layers employed in the multilayer elements for analysis of liquids 20 described in U.S. Patent 3,992,158. However, the transport of complex, high molecular weight substances, for example, proteinaceous substances having a molecular weight higher than albumin (which has a M.W. of about 60,000), has typically been carried out in conventional, non-fibrous, particulate 25 spreading layer structures only with dirficulty, typically exhibiting some chromatographing problems (sometimes refer-red to as "ringing") or requiring extended "spread" times (i.e., the time required ror the structure to take up, distribute within itself, and transport an applied liquid 30 sample) on the order of a minute or more.
The elements of the invention have a particulate structure comprising a plurality Or heat-stable, organo-polymeric particles non-swellable in and impermeable to the liquid under analysis and an adhésive ~or these particles 35 comprising an organic pol~mer dl~ferent from that of the particles. The adhesive is concentrated on the surface of the heat-stable particles in areas contiguous to ad~acent particles and bonds the particles into a coherent, three-dimensional lattice that is non-swellahle ln the liquid . , . :

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under ana~ysis. This lattice contains ~nterconnected void spaces amon~ the partlcles represen~ing a total void volume of about 25 to 80 percent t3 provide transport Or ~he aqueous ¦
liqu~d and prererably to render the lattice isotroplcally porous. The organo-polymeric part~cles typically have a particle size OI~ from about 1.0 to 200 microns. The adheslve ~or these particles is insoluble in the llquid under analysis and is presen~ in the element in an amount less than 10 percent by weight, preferably from Q.l to less than 5 percent by lC weight~ ~aRed on the:weight ~f ~he particles.
Because the adheslve contained in the element is concentrated at particle surface areas contiguous to ad~acent particles, the three-dimensional lattlce structure of the ele~ent exhibits a high void volume which remains substan-15 tially rree from adhesive. Moreover, because the elementcontains a small amount o~ adhesive based on the weight Or or~ano-polymeric particles, there ls llttle or no excess adhesive available to clog an~ ~ill in the lnterconnected void spaces of the three-dimensional lattlce. The non-23 swellability and lmpermeability properties Or the organo-polymeric particles and the ~nsolubility property of the adhesive represent rurther important ~actors contributing to the retention Or high void volume and advantageous llquid analysis and transport properties provided by the element o~
25 the invention.
In an especially pre~erred embodiment Or the lnvention, analytlcal elements are provided ror the analysis an analyte contained ln an aqueous liquid sample. These elements comprise an lnteractive compositlon ~or detection 30 of the analyte ln the aqueous ~ample associated with a particulate structure, as descrlbed above, ln which the organo polymerlc particles are impermeable and non-swellable to water and the adhesive ls water insoluble. In these - elements, the interactive composltion ror analyte detectisn 35 ls associated wlth the parklculate ~tructure in a manner er~ective to provide rluid contact between the lnteractlve compositlon and the particulate structure. The interactlve compositlon can there~ore be ~resent ln the matrix Or the ~`'' ~ '`` . ' .~ .
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particulate structure; it can be located in a separate zone of the element in rluid contact with the particulate structure; or certain components o~ the inter-active composition for analyte detection can be located in the particulate structure and other components of the nteractive composition can be distributed in one or more separate zones of the element which are in fluid contact with the particulate structure. In the latter two cases, an analytical element is provided which represents a 13 multi-zone element having at least one zone comprlsing the aforementioned particulate structure in fluid contact with at least one other zone comprising a separate reagent zone containing one or more components of an interactive composi-tion for analyte detection.
In a further embodiment, interactive compositions use~ul ~or the detection o~ various analytes present in liquids are affixed to the surface o~ the organo-polymeric particles contained in the particulate structure o~ these elements. In a further aspect o~ this embodiment, the 20 organic polymer of the particles advantageously contains a repeating unit comprising a chemical group representing an active bondin~ site for chemical attachment Or an interactive composition.
In another embodiment o~ the lnvention, multi-zone 25 elements containing, in fluid contact, at least two zones having a particulate structure, as described above, are provided. In one preferred aspect of this embodiment, the structure o~ each such zone has a dif~erent void slze whereby large molecules are retained in one zone while smaller 3 molecules migrate into a zone having a smaller void size.
In another especially preferred embodiment, the invention provides an analytical element having one or more zones comprising the particulate structure described above as a layer carried on a support. A radiation-transmissive 3~ support is particularly pre~erred to enhance and facilitate determination o~ detectable changes occurring in these elements by use of various radlometric detection methods.
In ~urther embodiments, analytical elements are provided ~ ' .. .

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which may contain one or more zones comprising a par-ticulate structure as described abo~e and one or more separate func~ional zones permeable to the liquid under analysis, such as reagent zones~ registration zones, radiation-blocking zones, selectively permeable barrier zones,detectable species mi~ration inhibiting zones, conventional isotropically porous nonfibrous spreading zones and the like, as described in US Pat-ents 3,992,158, 4,~42,335, 4,066,403 and 4,166,093.
The individual zones of these mul~ione elements are preferably present as superposed layers in fluid con-tact with one another.
The aforementioned particulate structure and an element containing the same, as described in fur-ther detail hereinafter, can also provide effective analytical elements for immunoassays. Such immunoas-say elements represent a particularly useful embodi-ment of the invention. Immunoassays are typically employed for anmalysis of extremely low concentrations of analyte contained in a liquid sample. However, the interactive compositions used in the assays, for exam-ple, immunoreagents, such as antibodies, antigens and haptens, and various detectable species, sometimes referred to as labels, associated with these immuno-reagents, often represent large, complex molecular species. Thus, "dry chemistry" analytical elements for immunoassays must be able to transport these large, comple~ substances without impeding, blocking or otherwise interfering with the migration of these molecules through the element structure. The particu-late structure described herein is ideal for transport of such large molecules.
The elements of the invention can be used essentially for transport of a liquid. Preferably, however, these elements can also provide for analysis of an analyte contained in a liquid. Analysis of a , ~a~

liquid by the method of the invention comprises the steps of:
(a) contacting together the liquid and the element of the invention to interact the analyte, or a reac-tion product of the analyte, for example, physi-cally or chemically, with the element to produce a detectable change within the element, and (b) detecting this change~ such as by an appropriate radiometric technique, to determine ~he presence and/or concentration of the desired analyte.
A further embodiment of the invention pro-vides a preferred method of making the above-described elements. This method comprises the steps of:
(a) forming in a liquid carrier a "stable dispersion"
of the organo-polymeric particles and the organic polymeric adhesive and (b) applying this dispersion to a support and removing the liquid carrier at a temperature below the heat-stability temperature of the organo-polymeric particles, such as by suitable drying conditions, to form, in situ, the desired particulate struc-ture of the invention.
Brief Description of the Invention Fig 1 îs a drawing illustrating, diagrammati-cally, the particulate structure comprising the organo-polymeric particle~ and adhesive contained in the elements of the invention.
Fig 2 is a black-and-whtie electron micro-graph obtained under 6000x magnification showing the three-dimensional lattice formed by the heat-stable particles and the adhesive in a preferred particulate structure of the invention. The electron micrograph shows the extensive void volume provided by the inter-connected void spaces of the lattice, as well as the concentration of the adhesive at those particle sur-face areas contiguous adjacent particles.

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Figs. 3 7 represent enlarged sectional views of certain preferred embodiments illustrating integral analyt-ical elements containing a zone having the particulate structure characteristic of the invention as a layer which, if desired, can be superposed over a support or other conti~uous zones in layer rOrm.
Fig. 8 represents an enlarged sectional view of a further embodiment illustrating a multi-zone analytical element having at least two adjacent abutting zones, at 10 least one of these zones containing the particulate structure characteristic of the present invent~on.
Fig. 9 represents an enlarged sectional view o~
another embodiment wherein an element o~ the invention contains at least two zones spaced apart from one another 15 until the time of use of the element, at least one of these 7ones containing the particulate structure characteristic Or the present invention.
Figs. 10-14 represent enlarged sectional views of various analytical elements particularly suited for 20 immunoassaY-DescriDtion Or Pre~erred Embodiments Particulate Structure An essential feature of the invention is the coherent, three-dimensional lattice formed by the organo-25 polymeric particles and the adhesive ~or these particles.The interconnected void spaces existing among the ad~acent particles of this lattice structure are essential to provide ~or transport of liquids and for substances, e.g., high molecular weight analytes, which may be contained in a 3liquid or introduced into the liquid as it is transported through the structure. Maintaining particulate integrity of the organo-polymeric particles in the lattice structure prevents the coalescence and flow of these materials into these void spaces; and the concentration o~ adhesive at 35those particle surface areas o~ the lattice which are contiguous to ad~acent particles insures that the adhesive does not flow into and clog these spaces.

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-A large portion of the particle surface area present in the lattice structure is e~fectively ~ree from adhesive. By retaining substantial portions of the particu-late surface area effectively free from adhesive, not only is "open space fill-in" within the lattice avoided, but also these surface areas are available for use as binding sites or equivalent "fixing" sites, for any of a variety of interactive compositions useful in a particular analysis.
The concentration of the adhesive at particle 10 surface areas contiguous to adjacent particles also prevents the adhesive from interfering with interactive compositions which may be fixed to these particle binding sites. Thus, if desired, various interactive compositions can be "pre-attached" to the surface of the organo-polymeric particles 15 without fear that large portions of the interactive composi-tion will be effectively overcoated, covered up, or other-wise inactivated by the adhesive contained in the particu-late structure.
The void spaces in the lattice structure represent 20 a void volume within the range of from about 25 to 80 percent. Typically, where it is desired to maximize void volume 7 preferred elements contain particulate structures having void volumes within the range of from about 40 to 80 percent. The presence of these interconnected void spaces 25 provide effective fluid flowpaths through the particulate lattice structure and, preferably, renders the lattice isotropically porous.
The term "isotropically porous~' and similar terms refer to porosity in all directions within the particulate 3 structure. The degree of porosity can vary, o~ course, depending on void size, void volume or other parameters.
Although the size of these void spaces is com-paratively large on a molecular scale and therefore capable of handling large complex molecular substances without 35 clogging, the absolute size of these void spaces is still small. Typically, the effective mean void size exhibited , , by these particulate structures is within the range of from about 0.1 to lx the mean particle si~e Or the organo-polymeric particles contained in the structure. Thus, liquid transport is facilitated by the capillary action Or the liquid bein~ drawn through these interconnected spaces within the particulate s~ructure Or the element. Stated in other words, the interconnected void spaces in these struc~
tures represent interconnected microvoids ror liquid trans~
port.
lC The size of the void spaces and the void volume of the particulate structure can vary widely and will depend upon a number o~ ~actors including the size of the particles contained in the structure, the method of preparing the structure, and the like. ~urther detail concerning these 15 various parameters is presented hereinarter. For any given element, the desired size of these void spaces will depend upon the particular liquid to be transported, i.e., its viscosity, and upon the size and molecular configuration of various components contained in the liquid or introduced 20 into the liquid as interactive compositions. The size Or void spaces in the particulate structures can be measured by conventional techniques such as mercury intrusion techniques.
Void volume can be calculated with reasonable accuracy by a variety of techniques such as described in Chalkley, Journal 25 of the National Cancer Institute, 49 p. 47 (1943) and by direct weighing and determining the ratio of actual ~eight of the structure to the weight o~ solid material equal in volume to that of the structure, comparably composed Or constituents from the structure.
3 A further advantageous reature of the particulate structure is its "metering" capability. That is, like the conventional particulate spreading compositlons described in U.S. Patent 3,992,158, these particulate structures, when ln a planar form, also can receive on one surface thereof, an 35 applied liquid sample and distribute the sample wlthin itself such that, at any given time a a uniform concentration .o~ the liquid sample and analyte contained therein is provided at the opposite sur~ace o~ the planar structure.

, It is possible to obtain such uniform concentrations over a range of sample volumes applied to the structure so that extremely precise sample application techniques are not required although approximate uniformity of applied sample volumes, e.g. ~ 10-20%, may be desirable to achieve pre-ferred spread times or the like.
The thickness, i.e., dry thickness, of the par-ticulate structure can vary widely depending upon the size of the organo-polymeric particles in the structure and the 10 specific use for which the structure is intended. For example, an element having a support bearing the particulate structure as a superposed layer typically employs a particu-late structure having a dry thickness within the range of from about 10 to about 500 microns. However, in certain 15 applications, structures having a thickness outside the aforementioned range may also be employed.
To provide further illustration of the particulate structure of the invention, Figs. 1 and 2 are attached.
Fig. 1 illustrates, diagrammatically, a preferred particulate 20 structure 3 as viewed under magnification containing an array of organo-polymeric particles 4 having an average particle size of from about 1 to 20 microns. The total amount of adhesive 5 contained in structure 3 of Fig. 1 is on the order of about 2 percent by weight based on the total 25 we`ight of adhesive 5 and particles 4 contained in structure 3. In Fig. 1 adhesive 5 is concentrated on particle surface areas 7 contiguous to ad~acent particles within structure 3, thereby forming the characteristic three-dimensional lattice structure of the invention. Other particle surface areas 3 may contain some amount of adhesive as indicated by particle surface areas 8 in Fig. 1, but the adhesive is concentrated ln particle sur~ace areas 7 contiguous to adjacent particles.
Therefore, large surface area portions of the ma~ority of particles 4 contained ln particulate structure 3 are effec-35 tively free from adhesive 5. Structure 3 contains a largenumber of interconnected void spaces 9 to provide llquid transport and to render the structure isotropically porous.
Fig. 2 ls a black-and-white electron micrograph showing an actual particulate structure 3 of the invention.

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The electron micrograph of Fig. 2 was taken at 6000x magni-fication. Adhesive 5 is visible ~n Fig. 2 concentrated at particle surface areas 7 contiguous to ad~acent particles.
Interconnected void spaces 9 are also readily visible in Fig. 2.
Heat-Stable Particles The organo-polymeric particles employed in the present invention are heat-stable particles. "Heat-stable particles" refers to particles which, upon exposure to 10 typical environmental temperature conditions, do not burst, become tacky, coalesce into an agglomerate of individual particles, or otherwise undergo signilicant physical altera-tion. If such physical alteration were to occur under these conditions, the void spaces in the particulate struc-15 ture of the element could be effectively plugged or seri-ously damaged, thereby impairing the liquid transport capability of the element. The "heat stability temperature"
of an organo-polymeric particle refers to the maximum temperature at which the particle can retain lts heat 20 stability properties. Therefore, the heat-stability temperature of an organo-polymeric particle typically corres-ponds to or is higher than the glass transition temperature of the polymer component of the particle. In general, particles are considered to possess sufficient heat stability 25 if they retain their initial physical shape and remain non-tacky at temperatures extending at least over the range offrom about 15C to 40~C, preferably from about 10C to 80C.
The particles used in the inventlon are also ~mpermeable and non-swellable in the particular liquid 30 intended for transport and~or analysis by the element.
These properties of the particles insure the structural integrity and retention of the void spaces within the element upon application of the liquid. As noted in the Related Art section, if the particles ~well in the presence 35 of liquid the void spaces of the element can become plugged.
- "Non-swellability" or "resistance to swell" re~ers to particles which exhibit little (i~e., less than about 20%, preferably less than 10% swell) or no swelling as determined by a swellability test. With respect to aqueous liquids, such a test can be carried out by forming a sel~-supporting film of the specific polymer under consideratlon for use as a particle material or a layer of the polymer9 such layer havin~ a dry thickness of from about 50 to 200 microns 9 on a suitable support, for example, a polystyrene rilm support, and evaluating the swell properties of this rilm or layer ln the presence of the desired liquid by use o~ a swellometer of the type described in A. Green and G. I. P. Levenson, Journal of Photo~raphic Science, 20, 205 (1972). Using this swellometer, the swell properties of the ~ilm or layer can be measured by determining the percent increase in the film or layer thickness which results from immersing the dry ~ilm or layer into a liquid bath at 38C for 2.5 minutes.
Although shape and size of the organo-polymeric particles can vary widely, in a preferred embodlment these particles are of substantially uniform size. Typically, the particles have a curvilinear surface and most preferably the particles are substantially spherical beads. Generally, these particles have a particle size within the range of from about 1.0 to about 200 microns.
The si~e of the organo-polymeric particles regu-lates, to an extent, the size of the void spaces contained in the particulate structure of the element. In a preferred embodiment, wherein the particulate structure o~ the element is intended for the transport Or aqueous liquids containing complete cellular structures such as red blood cells which can attain a size range of from about 6 to 8 microns, one would select large size particles for use in the elements of the lnvention. In such case, particle sizes within the 3 range of from about 20 to 200 microns, preferably about 20 to 100 microns can be employed.
In the case where one is concerned with the transport of large, complex molecules, such as macromolecules o~ biological origin, for example, llpoproteins, antlgens, and the like, somewhat smaller~ although still relatively large particles having a size range on the order of from about 2 to 20 microns, preferably about 2 to 10 microns can be emplo~ed. In the case Or aqueous flulds containing . . . . . ...................... . . .

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analytes of still smaller molecular size, for example, glucose molecules and the like, one can employ partlcles having a size within the range of from about 1.0 to 5 microns.
As noted, an especially pre~erred embodiment is an element having a particulate structure capable of transport-ing aqueous liquids containing large complex molecular substances; and accordingly, elements containing particulate structures employing particle sizes within the range of from about 2 to 200 microns represent an especially preferred and advantageous embodiment of the invention.
Based on thè organo-polymeric composition and the properties of impermeability and non-swellability exhibited by the particles employed in the invention, these particles are also insoluble in the liquid under analysis. Insolu-bility o~ a speci~lc material refers herein to insolubility of the material in the particular liquid of interest, e.g~, water, as measured at 20C and, in the case of water, at a pH of about 7Ø The particles employed in the invention 20 are typically and preferably solid (~.e., not hollow) particles. This is not an essential requirement, however, providing the particles have adequate heat-stability as discussed above.
The organo-polymeric composition of the particles 25 can be composed of a wide variety of organic polymers, including both natural and synthetic polymers having the requisite impermeability and non-swellability properties.
Because the elements can be employed to analyze a variety of liquids which can have widely varying properties, the 3 particular organo-polymeric composition Or these particles should be selected to match the particular liquld for which a specific element is intended. Thus, the organo-polymeric particles selected need not be impermeable and non-swellable in all liquids, but only that liquid for whlch an element 35 containing such organo-polymeric particles is intended for use. Such organo polymers can be thermosetting, thermoset or thermoplastic polymers. The polymers can be addition polymers or condensation polymers, such as polyesters, "` , : . ' ": . : ' : , :
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. ~. , -16a-polycarbonates, polyamides, sllicone polymers, etc. Pre~er-ably, the or~ano-polymeric particles are composed of addition polymers, including addition homopolymers and addition copolymers prepared from 2 or more addition polymerizable monomers. Especially preferred ln accord with one embodiment of the invention are addition copolymers prepared from a blend of two or more different addition polymerizable monomers.
Both the organo polymers Or the heat-stable 10 particles and the adhesive polymers described hereinafter in the Adhesive Section can be prepared by any of a variety of conventional polymerization methods. Typical addition polymerization methods include: solution polymerization (followed by an appropriate precipitation procedure in 15 the case of polymers formed into heat-stable particles), suspension polymerization (sometimes called bead polymeriza-tion), emulsion polymerization, dispersion polymerization, and precipitation polymerization. Condensation polymers used in the preparation of the heat-stable particles and 20 the polymeric adhesive can be prepared by conventional condensation polymerization processes, e.g., bulk and hot-melt polymerization.
In an especially preferred embodiment, the partic-ular organo polymer selected contains one or more reaction 3o ,;
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sites to link various interact1ve compositions to the sur~ace of the particles. This embodiment is particularly useful wherein the element of the invention contains an interactive composlt~on within the particulate s~ructure ~or use in an analytical reaction scheme used to detect ~he analyte of interest. Of course, where the element of the invention is employed solely as an aqueous transport struc-ture, or where a particular interactive composition con-tained in the structure is fixed within the structure by 10 physical means such as adsorption to the particle surface, or where it is unnecessary to fix the interactive composi-tion within the particulate structure, organo polymers without the above-described reaction sites can readily be employed in these particles.
1~ In accord with an especially preferred embodiment wherein the organo polymeric particles selected must be water impermeable and water-nonswellable, a partlal listing of representative addition homopolymers and copolymers useful for the organo-polymeric particles include polymers 20 prepared from an addition polymerizable blend of monomers.
Particularly, useful addition polymer~zable blends of monomers are blends wherein the total monomer composition of the blends have the following composition:
a. from 0 to 100, preferably 0 to about 99, weight 2~ percent of a polymerizable~ amino-substituent-free styrene monomer includlng derivatives and equivalents thereof, for e~ample, a styrene monomer having the following formula CHRl= CR2~/R3 wherein each of Rl and R2, which may be the same or different, represents a non-lnterferin~ sub-stituent such as hydrogen, halo7 or a substituted or unsubstituted, amino-free alkyl or aryl group having 1 to about 10 carbon atoms and R3 represents a non-interfering substituent such as hydrogen, halo, or a substltuted or unsubstituted, amino-free allphatic or aromatic group having 1 to about , : ~
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10 carbon a~oms, e.g., alkyl, alkoxy, aryl, or aryloxy group. Typical o~ such styrene monomers are styrene, vinyltoluene, t-butylstyrene, and equivalents thereof.
b. from 0 to about 25 weight percent of a polymeriz-able acrylic ester including derivatives and equivalents thereof, such as an acrylic ester having the formula CHRl=CH-COOR
wherein Rl is as defined above and R4 represents a hydrocarbyl group having from 1 to about 10 carbon atoms including hydrocarbyl groups such as aryl groups, alkyl groups, alkaryl groups, aralkyl groups, e.g., benzyl groups, and the like.
c. from 0 to 100, preferably O to about 75, weight percent of a polymerizable methacrylic ester including derivatives and equivalents thereof, such as a methacrylic ester having the formula CHRl=C - COOR

wherein R and R are as defined above.
d. from 0 to about 30 weight percent of a car-boxylic acid containing one or more polymerizable ethylenically unsaturated groups, such as meth-acrylic acld, acrylic acid, crotonic acid, ita-conic acid, and equivalents thereof;
e. from 0 to about 75 weight percent of a nitrile containing one or more polymerizable ethylenically unsaturated groups, such as acrylonitrile, meth-acrylonitrile, and equivalents;
3o ~. from 0 to about 20 weight percent of a polymerizable amine-substituted styrene monomer, includlng styrene monomers having N alkyl substltuted amino substituents on the phenyl ring of the styrene monomer~ such amine-substltuted styrene monomers - . . :
: , typically having the formula CHRl=CR~
(A n R6 ~
Am wherein each of n and p, which can be the same or different, represent 0 or 17 Rl-R3 are as deflned above, R6 represents an alkylene gr~up having rrom 1 to about 6 carbon atoms, and Am represents a primary, secondary, or tertiary amino group.
Typical Or such amine-substituted styrene monomers are Ngl~-dimethyl-r~-vlnylbenzylamine and styrenes containing i-alkyl substituted amino substituents, such as N-methylaminoethylstyrene and ~ -dimethyl-aminoethylstyrene.
g. from 0 to about 20 weight percent, preferably 0 to about 10 weight percent, of an addltion poly-1~ merizable monomer containing a crosslinkable group, including (1) addition polymerizable monomers which can be crosslinked by conventional gelatin hardeners, for example, aldehyde hardeners, haloethyl-sulfonyl hardeners, bis(vinylsulfonyl) hardeners, and the like. Particularly preferred such monomers which can be cross-linked by conventional gelatin hardeners are addition polymerizable monomers containing an active methylene group as described in ~.S. Patents 3,459~790;
3,488,708; 3,554,987; 3,~58,878; 3,929,482;
and 3,939,130; and (2) addition polymerizable monomers which can be 3 crosslinked by diamines~ such monomers containing a conventional gelatin hardening group, for example, aldehyde group-containing monomer~, haloethylsulfonyl group-containing :: ,, ,, ., - ~,. , - "
: ., : . ~ .. :. - , ~: . . ." . ' - ,, . . . " . . -,: - . . .. .... : . ..

: ., ,: ~ .

monomers, vinylsulfonyl group-containing monomers, and the like;
h. from 0 to about 20 weight percent of a polymeri-zable tertiary aminoalkyl acrylate or methacrylate monomer and equivalents thereof, such as dimethyl-aminoethyl acrylate, diethylaminoethyl methacryl-ate, and the like;
i. from 0 to 100, preferably 0 to about 75, weight percent Or a polymerizable, N-heterocyclic vinyl monomer and equivalents thereof, such as 4-vinyl-~ridine, 2-vinylpyridine, and the like;
j. from 0 to about 20 weight percent of a polymeri7-able acrylamide or methacrylamide monomer and equivalents thereof, including monomers having the formula o CHRl=CR7C Am wherein Rl and Am are as defined above and R7 represents hydrogen or methyl. Typical Or such acrylamide or methacrylamide monomers are N-isopropylacrylamide or N,N-dialkylacrylamide or N,N-dialkylmethacrylamide; and k. from 0 to about 20 weight percent, preferably 0 to about 5 weight percent, Or a crosslinking monomer containing at least two addition polymerizable groups, such as divinylbenzene, N,N-methylenebis-(acrylamide), ethylene diacrylate, ethylene di-methacrylate, and equivalents thereof.
It is understood, of course, that the above-noted monomer blend compositions that contain 100 weight percent of a single monomer result in addition homopolymers.
A partial listing of representative polymers ~or use in making the organo-polymeric partlcles is set forth ~n 35 Table I. The numbers in brackets following each of the polymer names represents the weight ratlo of monomers con-tained in the monomer blend from which the polymers are polymerized.

i: - . . .

.- : . .

TABLE
1 . Polystyrene 2. Poly(styrene-co-methacrylic acid) [98/2]
3. Poly(vinyltoluene-co-~-t-butylstyrene-co-methacrylic acid) [61/37/2] Tg = 100 C

4. Poly(vinyltoluene-co-p-t-butylstyrene co-methacrylic acid-co-divinylbenzene) [60/37~2/1]

5. Poly(methyl methacrylate) ; 6. Poly(styrene-co-vinylbenzyl chloride-co-methacrylic 10 acid) [78/20/2] Tg = 1~3 C
7. Poly(styrene-co-N,N,N-trimethyl-N-vinylbenzyl-ammonium chloride-co-methacrylic acid) t88/10/2]
8. Poly(styrene-co-divinylbenzene) ~98/2]
9. Poly(styrene-co-butyl acrylate-co-methacrylic ac~d) [88/10/2] Tg = 84 C
10. Poly(styrene-co-methacrylic acid-co-dlvinylbenzene) [70/25/5 and 98/1/1]
11. Poly(vinylbenzyl chloride-co-methacrylic acld-co-divinylbenzene) [g3/2/5] Tg = 68C
20 12. Poly(styrene-co-2~hydroxyethyl methacrylate-co-methacrylic acid) [88/10/2] Tg = 99C
13. Poly(methyl methacrylate-co-butyl acrylate) ~70/30] Tg = 70 C
14. Poly(styrene-co-acrylonitrile) [70/30] Tg = lQ~C
15. Poly(methyl methacrylate-co-N-(_- and ~-vinylbenzyl)-N,N-dimethylamine hydrochloride-co-ethylene dimethacry-late) [70/20/10~
16. Poly(methyl methacrylate-co-2-(N,N~diethylamino)-ethyl methacrylate hydrochloride-co-ethylene dimethacrylate) ~70/20/10]
The partlcles typically comprise at least about 25 weight percent and preferably 50 weight percent or more Or the above-described organo-polymeric composition. In m ny - embodiments3 these particles are composed entlrely, i.e.~
100 welght percent, Or such organo-polymeric materlal. ~he 3~ remalnder Or these partlcles can be composed o~ other addenda, - ~or example, colorants such as plgments or dyes; radiation-blocking agents includlng colorants and other opacirying ... .
.
.
.: , addenda; fluors; fillers; magnetic addenda, e.g., magnetite;
and the like, provided the requisite impermeability and non-swellability properties of the particles are maintained, and the addenda do not inter~ere with the analysis to be carried out in the element in which the addenda is incor-porated. Broadly~ such addenda can be described as noninterfering addenda and can be employed to enhance or facilitate a particular analytical procedure or test result.
Adhesive The adhesive employed in the invention bonds the organo-polymeric particles to one another to provide the coherent, three-dimensional lattice of the elements. The adhesive is composed of an organo polymer different from the specific polymer contained in the particles, although quite commonly the adhesive represents a polymer containing many repeating units which are identical or similar to some of those present in the polymer composition of the particles.
Within the context of the present specification, organo polymers useful for the heat-stable particles are considered different from those useful as adhesives provided they have differing viscosities and possess appropriate heat-stability or glass transition temperatures, even though they may be composed of identical repeating units.
In accord with an especially preferred embodiment, both the adhesive and the organo-polymeric particles repre-sent an addition polymer with the adhesive representing an addition copolymer of two or more different addition polymer-izable monomers, at least one of the addition polymerizable monomers of the adhesive being common to one of the monomers 3 of the organo-polymeric particles.
The adhesive represents a polymer which, when incorporated in the particulate structure of the element, '.' ; ' ' .
.

is insoluble in the liquid to be analyzed or transported by the element. Thus, suitable adheslves include polymers that are initially soluble in the liquid but become insoluble during formation of the particulate structure, for example, by crosslinking. Preferably the adhesive is also non-swellable in the liquid. Howe~er, because of the small amount of the adhesive contained in the particulate struc-ture of these elements, namely less than 10 weight percent, preferably from about 1 to less than 5 weight percent, based on the weight of the particles contained in the structure, non-swellability of the adhesive, although prererred, is not essential.
The small amount o~ adhesive in the particulate structure is an important factor contributing to the desired retention of void spaces within the structure which are substantially free ~rom and unclogged by adhesive. In accord with certain preferred embodiments, the amount of adhesive contained in the particulate structure represents from about 2 to 4.0 percent by weight based on the dry weight of the particles in the structure.
Preferred water-insoluble adhes~ves for use in the invention are addition homopolymers and copolymers, particu-larly addltion copolymers, prepared from an addition polymer-izable blend of monomers selected from the ~ollowlng group:
A. a monomer blend containing from about 1 to 35 weight percent, preferably about 10 to 30 weight percent o~ one or more polymerizable styrene monomers as defined in (a) above with the re-mainder of the blend comprising addition 3 polymerizable monomers selected from the group consistlng of alkyl acrylates or methacrylates and mixtures thereof wherein the alkyl group of these acrylates and methacrylates prererably has from 1 up to about 6 carbon atoms, such as n-butyl acrylate, n-butyl methacrylate, ethyl acrylate, - and the llke;
B. a monomer blend containing from about 20 to 95 weight percent, preferably 50 to 95 weight percent of monomers selected from groups (a), (b)~ (c)~

~ .

(gl), (g2), and (k), preferably groups (a)-(c) and (k) noted above such as styrene, ethyl acrylate, n-butyl acrylate, n-butyl methacrylate, 2-ethyl-hexyl acrylate, and methyl acrylate, with the remainder of the monomer blend comprlsing one or more addition polymerizable monomers having an active hydrogen or salts thereof. ~he term active hydrogen is defined in accord with the derinition set forth by J. March, "Advanced Organic Chemistry:
Reactions, ~lechanisms, and Structure~" ~IcGraw, Hill, Inc., page 471 (1968) which de~ines active hydrogen as one whlch will react with methyl magnesium bromide, i.e., as in the Zerewittenoff Process. A partial listing of representative addition polymerizable monomers containing active hydrogen or salts thereof includes acrylic acid;
methacrylic acid; vinylbenzyl alcohol; hydroxy-alkyl acrylates and methacrylates having ~rom 1 to about 6 carbon atoms in the alkyl group thereof;
and an addition polymerizable, sulfo- or sulfate-substituted monomer, including sulroalkyl acrylates or methacrylates such as N-sulfoalkylacrylamides or N-sulfoalkylmethacrylamides, such as 2-methyl-2 acrylamidopropane sulfonic acid, as well as the alkali metal and ammonium salts thereof, and other addition polymerizable alkyl sulfonate monomers, aryl sulfonate monomers, e. g., 4-sulrostyrene, alkyl sulfate monomers, aryl sul~ate monomers, and equivalents thereof (a partial listing of repre-3 sentative speclfic addition polymerizable, sulro-or sulfate-substituted monomers may be found in the following U.S.Patents: U.S. 2, 923,734;
3,024,221; 3,265,654; 3,277,056; 3,~11,911;
3,5069707; 3,525,768; and 3,547,899); addition polymerizable monomers as described in groups (gl) and t g2) above; acrylates and methacrylates o~
poly(alkylenediols) such as poly(ethylene glycol), ~or example, an acryllc ester Or Tergitol 15-s-12, a poly(ethylene glycol) ether of a linear , .

.' ' , secondary alcohol sold by Union Carbide Corp. and alkali metal and ammonium salts of the foregoing monomers capable of such salt rormation. Pre~-ferred active-hydrogen containing monomers or salts thereof include acrylic acid, methacryllc acid, 2-acrylamldo-2-methylpropanesulfonlc acld, and the alkali metal and ammonium salts of these acids; and C. a monomer blend containing rrom about 15 to 100 weight percent of one or more monomers selected from the group conslsting Or l-vinylimidazole, vlnylbenzyl alcohol, ethyl acrylate, or an acryl-amide or a methacrylamide such as N-isopropyl-acrylamide with the remaining monomers o~ the blend comprising addition polymerizable monomers as described in group (gl) monomers above such as 2-acetoacetoY~yethyl methacrylate.
It is understood, Or course, that the above-noted monomer blends in group (C) that contain 100 weight percent 20 Or a single monomer result in addition homopolymers.
Table II sets ~orth a partial listing of repre-sentative polymers of surricient water-insolubility to be useful as water-insoluble adhesives in the structure of the present invention. The numbers in brackets following each 25 O~ the polymer names represents the weight ratlo of monomers s contained in the monomer blend from which the polymers are polymerized.

3o TABI.~ II
1. Poly(n-butyl acrylate-co styrene-co-2 acrylamido-2-methylpropanesul~onlc acid) t70/20/10~ Tg - -15C.
2. Poly(bu~yl acrylate-co-styrene~co-2 acryl-amldo-2-methylpropanesulfonic acid) ~76/21/3] Tg = -18C.
3. Poly(ethy~ acrylate~co-acrylic acid-co~2-acetoacetoxyethyl methacryla~e-co-2-acryl-amido-2-methylpropanesulfonlc acid) in the following welght ratios: a) [71/24/4~1 Tg = -25C. and b) ~67J16~16/1 4. Poly(vinylbenzyl alcohol) 5. Poly(ethyl acrylate)

6. Poly(N-lsopropylacrylamlde)

7. Poly(2-hydroxyethyl methacrylate-co-2-aaetoacetoxyethyl meth crylate) [15/85] Tg = -20C.

8. Poly(n-butyl acrylate-co-acrylic acld) [75/25]

9. Poly(n-butyl acrylate-co~acryllc acld-co-methacrylic acld-co-ethyl acryloylacetate) [70/5/1~/10]

10. Poly(n-butyl acrylate-co-acryllc acid-co~
ethyl acryloylacetate) [75/15/10~

11. Poly(n-butyl acrylate-co-methacrylic acid-co-2-acetoacetoxyethyl methacrylate) [56/34~10~ Tg = -16C.

12. Poly(n-butyl acrylate-co-styrene) [70/30]

13. Poly(n-butyl acrylate-co-2~acrylamido-2-methylpropanesul~onic acid-co 2-aceto-acetoxyethyl methacrylate) ~B5/10/5~ Tg ~ -44G.

14. Poly(n-butyl acrylate-co-acryllc acid~co 2_ acetoacetoxyethyl ~ethacrylate-co-2~acryl-~mids-2-methylpropanesul~onlc acid) t67/16~16/1~ Tg 2~ -18C.

15. Poly(n-butyl methacrylate-co-styr~ne~
~0/10]

; -27-

16. Poly(ethyl acrylate-co-styrene) t70/30~

17. Poly~n-butyl acrylate~-co-2-acrylamido-2-methylpropanesul~onic acld) ~90/10] Tg = -46C.

18. Poly(n-butyl acryla~e-co-styrene) ~50/50]

19. Poly(2-ethylhexyl acrylate-co-acrylic acld-co-2~acetoaceto~yethyl methacrylate-.
co-2-acrylamido-2-methylpropanesul~onic acid) [67/16/16/1] Tg = -33C.

20. Poly(n butyl acrylate-co-methacryllc acid) [70/30]

21. Poly(ethyl acrylate-co-acrylic acid):
a) [80/20] T~ s 10C. and b) ~70/30~

22. Poly(butyl acrylate~co~styrene-co-2-aceto-acetoxyethyl methacrylate-co-2-acrylamido~
2-methylpropanesulronlc acld) [70/22/6/2]

23. Poly(butyl acrylate-co-s~yrene-co-2-acrylamido-2-methylpropanesulfon~c acld-co-dlvinylbenzene) ~69/20/10/1]

24. Poly(acrylamide-co-2-acetoacetoxyethyl methacrylate):
a) [20/80] Tg = 20C. (possibly crosslinked) and b) [15/85~

. . :

"'`, " "' .

Typically, the adhesive polymers have a glass transition temperature, Tg, which is at least 20C., pre-ferably 30C., less than the heat-stability temperature of the organo polymers contained in the heat-stable particles.
Preferred polymeric adhesives have a glass transition temperature below about 80C, typically less than 30C (as measured under high relative humidity conditions ~80% ~H).
Adhesives having such a glass transition temperature can easily be rendered flowable without affecting the heat stability of the organo-polymeric particles with which they are combined. The term glass transition temperature is defined herein to be that temperature at which the polymer changes from a glassy polymer to a rubbery or flowable polymer. Glass transition temperatures Or polymers des-15 cribed herein can be measured, for example, as described in"Techniques and Methods of Polymer Evaluation", Vol. 1, Marcel Dekker, Inc., N.Y. (1966).
Preparation of Particulate Structure Various methods may be employed for preparing 20 these particulate structures. In accord with a preferred embodiment, an especially useful method of making these structures comprises:
(a) forming in a liquid carrier a "stable dispersioni' of the organo-polymeric particles and the organic polymer ~5 adhesive, and (b) applying this dispersion to a support and removing the liquid carrier at a temperature below the heat-stability temperature of the organo-polymeric particles, such as by suitable drying conditions. The organo-polymeric particles 3 are dispersed in the aforementioned "stable dispersion" to retain their particulate integrity. The organic polymer adhesive can be dispersed or dissolved in the liquld carrier vehicle of the stable dispersions. When the organic polymer adhesive is dispersed in the stable dispersion, the liquid 35 carrier is preferably removed from the dispersion (following its application to the support) at a temperature abo~e the glass transition temperature of the polymeric adhesive but below the heat-stability temperature of the organo-polymeric particles.

' - , , Tne term "stable dispersion" is defined to mean that the particles and the adhesive remain admixed in the carrier without forming an agglomerated mass of particles and adhesive. Dispersions useful in preparing the particu--late structure need not remain stable for extended periodsof time, but should re~ain as a stable dispersion ~or a time sufficient to apply the dispersion to a substra~e serving as a temporary or permanent support ror the resultant particu-late structure. If the particles or adhesive settle out 10 of the dispersion, the adhesive or particles may be re-dispersed by agitating the dispersion.
To accomplish the formation of such stable dis-persions, a wide variety o~ techniques can be used. A
partial llsting of representative techniques is described 15 briefly herein. These techniques can be used individually or in combination. Of course, this listing of useful tech-niques is not exhaustive~ and therefore other techniques ~or formation of a stable dispersion can also be employed in step (a) Or the preferred method of making these partlculate 20 structures without departing from the spirit or scope of the invention. One useful technique is the addition of a surfactant to the liquid carrier to facilitate distribution and stabilization of the particles or the adheslve ln the dispersion and prevent rapid agglomeration and settling out

25 of these components. A partial listing of representative surfactants which can be employed includes non-ionic sur~ac-tants such as Zonyl FSN, a fluorochemical from duPont;
Triton X-100, an octylphenoxy polyethoxyethanol from Rohm and Haas; and Surfactant 10G , a nonylphenoxypolyglycidol 3 from Olin Corp.
In addition to the use of surfactants, formation of stable dispersions of the organo-polymeric particles and adheslves can be facilitated by controlling the order of addition of the adhesive and the particles to the liquid 35 carrier. For example, stable dispersions of certain organo-- polymeric particles and adhesives, which are normally , , di~ficult to form, can be formed by first combining the particles and the liquid carrier followed by addition of the adhesive; in other cases, depending upon the partlcular organo-polymeric particles and adhesive, stable dispersions may be achieved by ~irst combinlng the adhesive and the liquid carrier followed by addition of the partlcles.
Controlling the rate o~ addition of the organo-polymeric particles or the adhesive to the liquid car~ier can also facilitate obtaining stable dispersions. For lQexample, certain particles and adhesives, which normally do not form stable dispersions, can be formed into stable dispersions by adding only a portion Or the total amount o~
particles to be included in the dispersion together with the adhesive, followed thereafter by the addition of the remain-1~ ing amount of the particles. Similarly, this technique canbe used to control the rate of addition o~ the adhes~ve by adding the adhesive in incremental amounts to a liquid carrier which already contains the total amount of the particles.
2~ Still other techniques which can be employed to produce stable dispersions include procedures such as sonication treatments, physical blending and agitatlon treatments, pH ad~ustments, and the like.
In an especially preferred embodiment, format~on 2~ of a stable dispersion is facilltated by matchlng the specific gravity of the organo-polymeric particles and that o~ the carrier liquid. When the specific gravity of the particles is matched to that o~ a particular carrier liquid, such particles are often referred to as "neutral buoyancy"
3 particles. By use Or neutral buoyancy particles 3 one can reduce or eliminate the problem whereby certain otherwise useful particles are so dense that they immediately settle out o~ the dispersion, rather than belng distributed through-out the liquld carrier.
3~ Neutral buoyancy particles can be prepared by regulating the polymerization process for the organo polymer of a particular particle composition to obtaln an organo polymer havlng a predetermined specl~lc gravity relatlve to a desired carrier liquid. Alternatlvely, various rillers .

. . ; .

,~ . . .

can be blended with a particular organo polymer selected for use to obtain a resultant particle composition of polymer and filler having a bulk speclfic density similar to that of the desired carrier liquid.
In addition, one can match the specific gravity of the carrier liquid to that of the organo-polymeric particles by selecting a carrier liquid for use which has a specific gravity similar to that o~ the particles. In general, good results have been obtained in the method of the invention by ~'selecting carrier liquid compositions and particle compositions having a specific gravity within the range of from about 0.7 to 1.3.
When the particles have a specific gravity above about 1.0, it may be desirable to introduce a viscosity 1~ modifying agent into the carrier to obtain a stable dispersion.
This can be done, for example, by addition Or polymeric viscosity modifying agents such as hydroxyethyl cellulose, carboxyethyl cellulose, or derivatives thereof.
In another especially prererred embodiment, the 2C stable dispersion of the particles and the adhesive is ~ormed using an adhesive prepared as a latex. In one such embodiment, the adhesive is separately prepared as an aqueous latex, the latex comprising the adhesive polymer and any desirable or necessary surfactants as a finely-divided 2; discontinuous phase in an aqueous liquid vehicle as a con-tinuous phase, and then the organo-polymeric particles are admixed with the latex. In such case, the continuous phase o~ the latex, e.g., the aqueous vehicle, serves as at least a portion of the liquid carrier Or the stable dispersion Or 30 adhesive and organo-polymeric particles. This embodiment advantageously ~acilitates the maintenance of the adhesive in finely-divided, discrete form within the dispersion o~
adhesive and particles. This is desirable because it reduces undesired coalescence and agglomeration of the adhesive and the particles in the dispersion. In addition, it is belleved that use o~ the adhesive ln latex form promotes the concen-tration of the adhesive at discrete surface areas of the ..

,~ :
, L~ 6 4~ 4.

particles, as the liquid carrier is removed ~rom the particles and adhesive following application o~ the stable dispersion to a substrate. Moreover, use of the adhesive ~n latex form allows one to employ water-insoluble adhesives ln a dispersion containing an aqueous carrier.
When the adhesive is prepared as a latex9 these latexes typically contain an amount Or the polymer within the range Or from about 5 to 50 weight percent, based on the total wei~ht of the latex including the adhesive polymer, 10 the aqueous latex vehicle, and any necessary or desirable sur~actants. Such latexes can be prepared by a varlety of well-known latex techniques, such as those describedg for example, in C. E. Schildknech~, "Vinyl and Related Poly-mers", John Wiley & Sons, Inc., N.Y. (1952) and C. S.
15 ~arvel, "An Introduction to the Organic Chemistry of High Polymers, John Wiley & Sons, Inc., N.Y. (1959).
In general, stable dispersions o~ the organo-polymeric particles and adhesive in a liquid carrier vehicle contain from about 1 to 50 weight percent Or the particles 20 and from about 0.01 to 5 weight percent of the adhesive.
Typically, the temperature of the liquid carrier during rormation of the stable dispersion is at a level effective to maintain the organo-polymeric particles in a non-tacky state. That is~ the temperature is maintained at 25 a level below the heat stability temperature of these particles. This facilitates retaining the particulate in-tegrity of the particulate structure which is, Or course, hlghly desirable so that the void spaces o~ this structure remain open and unclogged.
3 Having ~ormed a stable dispersion in accord with step (a) Or the preferred method, step (b) is carrled out by applying the dispersion to a substrate and removing the carrier liquid. Typically~ this is accomplished by heating at a temperature below the heat-stabllity temperature o~ the 35 organo-polymeric particles. In the case where the adhesive - ls dispersed in the stable disperslon, the liquld carrier is pre~erably removed at a temperature which is above the ; ' glass transition temperature of the adhesive poly~er but below the heat-stability temperature Or the organo-polymeric particles. The polymeric adheslve thereby enters a ~lowable and tacky state. In this state, adhesion to the surface Or the organo-polymeric particles is facilitated. In addition, by placing the adhesive in a flowable state, one theoret~
ically can take advantage of the capillary pressures develop-ing between adjacent heat-stable particles as the structure is formed in situ during step (b) of the process. That is, 1~ it is believed that capillary pressure forces, which will become greatest in those regions of the structure wherein one particle is closely ad~acent to the surface Or another particle, can advantageously be used to draw the flowable adhesive to these regions and thereby enhance the concentra-1~ tion of the adhesive at those particle surface areas whlchare contiguous to adjacent particles.
Thus, ln step (b) Or the method, the stable dispersion is applied to a substrate, e.g., a temporary or permanent support, and the liquid carrier of the dispersion 2C is removed, such as by appropriate drying conditionsg to form, in situ, the desired three-dimensional particulate ~tructure. Typical drying conditions for removal of the liquid carrier are temperatures within a range of from about 10C to 65C. As step (b) is carried out, the adhesive ls 2~ concentrated at surface areas of the organo-polymeric particles contiguous to ad~acent particles and the adhesive bonds ad~acent particles together inko a coherent, partlculate structure. Care should be exercised throughout step (b) of the method to avoid exceeding the heat-stability temperature 3 of the organo-polymeric particles so that the resultant structure maintains i~s particulate integrity and retains the void spaces which are ~ormed among individual particles as the liquid carrier vehicle is removed.
The size of the void spaces obtained by the method 35 are influenced by a number of ~actors~ including, among others, size o~ the organo-polymeric parkicles, amount of adhesive and particles contained ln the stable suspension per unit volume, and rate of liquid carrier removal.

.
, ~ ~ "

Depending upon the nature of the adhesive i.e., whether it forms a suitable bond quickly or slowly or whether it requires further curing to achieve optimum bond strength, one can optionally provide a further heat treat-ment of the particulate structure to obtain optimum bondingand coherency of the structure. Again, of course, one should avoid using temperatures in this optional step which exceed the heat-stability temperature o~ the organo-polymeric particles.
The liquid carrier in which the organo-polymeric particles and adhesive are formed into a stable dispersion is typically an aqueous liquid, although other liquid carriers such as various organic liquids may also be employed provided the heat-stable particles are insoluble in the 1~ carrier so that their particulate character is retained. In one preferred embodiment as described above, the adhesive is also insoluble in the carrier so that it may be dispersed among heat-stable particles as a discrete discontinuous phase within the dispersion, thereby aiding the avoidance 20 of the formation of a substantial layer of the adhesive completely surrounding the particles. A partial listing of representative carrier liquids in addition to water, in~
cludes water miscible organic solvents, agueous mixtures of water and water miscible organic solvents, and suitable 25 water immiscible organic solvents. Typical water miscible organic solvents include lower alcohols, i.e., alcohols having 1 to about 4 carbon atoms in the alkyl group thereof;
acetones; and ethers such as tetrahydrofuran. Typical water immiscible solvents includes lower alkyl esters, e.g. ethyl 3 acetate, and halogenated organic solvents3 e.g., halogenated hydrocarbons.
_ teractive Compositions The particulate structures of the invention can advantageously contain one or more interactive compositlonsg 35 although the presence of such compositions in the particulate structure is not required. These compositions contain one or more active components that undergo lnteraction with an analyte, or a reaction or decomposition product of the analyte, or with each other upon application of a liquid .. ~ , . . .

sample containing the desired analyte to an analytical element including the particul~te structure. Such interaction can cause the release of a preformed detec-table species within the element, the formation of a detectable species or otherwise produce A detectable change in the elemen~. The term "interaction" is meant to refer to chemical activity, c~talytic activi~y as in the formation of an enzyme substrflte complex, immuno-genic activity as in an antigen-antibody reaction, and any other orm of electrical, chemical or physical interaction which can release, produce or otherwise provide within the element a detectable change which is directly or indirectly indicative of the presence and/or concentration of a desired ~nalyte, or a reac-tîon or decomposition product of the analyte.
Preferably (although note requîred), the detectable change which is produced is radiometrically detectable. Radiometric detection refers to detection by use of electromagnetic radiation-measuring tech-niques such as fluorimetry, colorimetry~ radioactive counting, phosphorimetry and the like.
As will be appreciated, ~mong the various com-ponents which can be present in interactive composi-tions are colorimetrically detectable dyes, pigments and complexes; fluorimetrlcally detectable dyes, pig-ments and complexes; phosphorescent tags; r~dioactive tags; chemical reagents; immunoreagents such as anti-gens, haptens, antibodies and antigen-antibody com-plexes; enzymes; and precursors an reaction products of the foregoing components. For further detail with respect to use of certain of these components, refer-ence may be made to US Patents 3,992,158 and 4,144,306 and Canadian Serial No 316,64~ filed November 14, 1978.
Although not required, the interactive compo-sitions, if ~n the particulate structure, can be immo-bilized ~o minimize or prevent undesired migration of the composition within the structure or other zones : , - ~ . ., :, . , ~ :,::

-36- ~
of an element containing the particulate structure. Immo-bilization can be erfected by a variety of means lncluding physical adsorption and chemical bonding to the heat-stable particles of the structure. For example, those heak-stable particles which are prepared from polymers containing an active linking or bonding site can advantageously be chemi-cally bonded to one or more components of a particular interactive composition by establishing a covalent bond between this site and a reactive group of the interac~ive lOcomponent. In addition to covalent bonding, ionic and hydrogen bonding can also be used where appropriate. In other cases, the molecular size or configuration Or the interactive composition may be effective to physically entrap and immobilize a particular interactive compositlon 1, in the particulate structure without use o~ any special physical adsorption or chemical fixing technique.
Element Structure The elements o~ the invention containing the above-described particulate structure can have any one of a 2~ variety of different configurations. Certain preferred embodiments illustrating representative configurations of the element are described hereinafter, but lt will be understood that other configurations o~ such elements although not specifically described are also consldered 2; within the scope of the invenkion.
In accord ~ith one embodiment, an element of the invention comprises the above-described particulate struc-ture optionally carried on a suitable support, preferably a radiation-transmissive support. In this configuration, 3 assuming no support is present, the element contains simply the above-described particulate structure. This particulate structure provides a remarkably erficient liquid transport means and can be ~ormed lnto a variety of shapes. In a 3~ typical embodiment, the structure is formed into a substan-tially coplanar con~iguration, ~or example, as a layer carried on a permanent or temporary support.
Fig. 3 illustrates a representative element of the invention lncluding the above-descrlbed particulate structure `', ~ ' , , ;, 3~

1 carried on a suitable support 2. Where structure 1 has sufficient durabil~ty or ln situations where durability requirements are not particularly critical or demanding, support 2 in Fig. 3 may be unnecessary. As will be apparent, the element Or Fig. 3 can be used simply to transport a liquid by contacting the liquid and the element together.
If desired, any one of a variety Or interactive compositions may be present within particulate structure 1 of Fig. 3 to carry out one or more interactions between or among various analytes contained in the liquid.
Fig. 4 illustrates another embodiment of an element containing at least two dirferent zones, zone 3 and 4, carried on a support 2. Each of zone 3 and zone 4 represent a particulate structure Or the invention. T~lo different zones 3 and 4 are present in the element of Fig. 4 to facilitate multiple treatments or operations on a particular liquid sample applied, for example, f~rst to zone 4 from which it is transported by the particulate structure Or zone 4 into zone 3 which is in fluid contact with zone 4. For - 20 example, the liquid sample can be sequentially exposed to two separate interactions or a series Or sequential inter-actions by incorporating different interactive compositions in each of zone 3 and zone 4. Alternatively, or in addi-tion, one can vary the average size of the organo-polymeric 25 particles employed in each of zone 3 and zone 4. For example, if zone 3 has a smaller pore size or average void space than does zone 4, one can effectively trap or remove components contained in a liquid which have a physical slze exceeding the pore size of zone 3 while permitting other 3 smaller components to be transported through zone 4 into zone 3. In this manner a multi-zone element such as illus-trated in Fig. 4 can be used to separate various liquid component~, based on their physical size, into two or more distinct zones of the element.
Fig. 5 illustrates yet a further embodiment o~ the element whereln the particulate structure is present as zone 1 of the element and zones 5 and 6 of the element represent , . ," ~ . .,"~

. .

other functional zones or layers. For example, zone 1 may be used as a ~spreading zone which meters and dis-tributes an applied liquid sample to a separate reagent zone 5 containing one or more components of an interac-tive composition which, upon interaction with an ana-lyte of the liquid, produces or releases a detectable product wi~hin the element. Such an element can optionally have present, as illustrated in Fig 5, one or more intermediate zones between spreading zone 1 and reagent zone 5. Such intermediate zones can serve as adhesive or subbin~ layers to improve adhesion between spreading zone 1 and reagent zone 5, or as a radiation-blocking zone to block or screen any undesired back~
ground color or other optical interferent of zone 1 from the detectable product released or formed in zone 5. The use of such radiation-blocking zones is further illustrated, for example, in the multilayer analytical elements described in US Patent 4,042,335. Or, inter-mediate zone 6 of Fig 5 may represent a detectable spe-cies migration-inhibiting layer such as described in US
Patent 4,166,093, to inhibit or prevent undesired back-migration of detectable species formed in reagent zone 5 into spreading zone 1 where the detectable species may become masked or otherwise difficult to detect.
Alternatively, intermediate zone 6 of Fig 5 can repre-sent a conventional nonfibrous isotropically porous spreading layer composed of, for example, a blushed polymer, a mixture of a blushed polymer and a particu-late material, or a mixture of a polymeric binder and microcrystalline cellulose particles. Each of the foregoing conventional spreading layer compositions is more specifically described in the aforementioned US
Patent 3,992,158.
More than one intermediate zone 6 may be pres-ent in a multizone element of the type illustrated in Fig 5. These zones may perform any of a wide variety , :: ,., ;
: , ' ~', .................................. :
,, -'. ~

of functions, only some of which have been described herein. For example, the multizone element illustrated in Fig 7 contains two intermediate zones 6 and 7, each of which can have any of a variety of functions. For example, each of zones 6 and 7 can be a subbing zone, a radiation-blocking zone, a detectable produc~
migration-inhibiting zone, a conventional nonfibrousg isotropically porous spreading zone, an additional rea-gent zone or an additional particulate structure-containing zone of the type described in the present invention, and the like.
Fig 6 represents another embodiment of the invention wherein the particulate structure is present as spreading zone 1 of the element. Reagent zone 5, as described above in Fig 5, is present. Also present in fluid contact with zones 1 an 5 is registration zone 10 to receive reaction products or detectable species released or formed in the element. Registration zones such as zone 10 of Fig 6 are further described in US
Patents 4,042,335 and 4,144,306.
Fig 8 illustrates yet another embodiment of an element containing a particulate structure as described herein. This element is also a multizone element, bu~
differs from previously illustrated multizone elements in that zones 1 an 5 of the element of Fig 8 are adja-cent abutting zones, such as a particulate structure spreading zone 1 and a reagent zone 5, rather ~han superposed layers as illustrated in elements of Fig 5 and 6. An optional support 2 is also shown in the ele-ment of Fig 8. Of course, as will be apparent, the element of Fig 8 can have other zones in addition to zones 1 an 5 illustrated in Fig 8, and each of these zones can represent a particulate structure of the invention, optionally having a different effective void or pore size. Or, the element can contain only one zone containing a particulate structure with each of ' the other optional zones having a different function and composition.
Fig 9 illustrates a further embodiment wherein an element contains at least two zones, e.g., zones 1 and 5 as described in Fig 5, optionally carried on sup-port 2, these zones intially being spaced apar~ by spacer means 9. Under conditions of use of the ele-ment, these zones are brought into contact such as by application of suitable compressive force to zone 1 of Fig 9 which causes pressure-deformable spacer means 9 in Fig 9 to deform and brings zone 1 into fluid contact with zone 5. Such a structure can be useful in an ana-lytical element where, for example, ~wo different interactive compositions are contained in zones 1 and 5 which would interact with one another prior to use of the element if zones 1 and 5 were maintained in physi-cal contact.
As indicated above, various optional func-tional zones (or layers) and supports can be present in the multizone elements of the invention. Such optional zones can be located adjacent the particulate structure of the invention or they can be superposed over or under the particulate structure. In addition to the specific functional zones discussed above, these optional zones can also include, among others, filter-ing zones to filter out or remove particular components of applied liquid samples, as described in US Patent 3,992,158; barrier compositions having a predetermined selective permeability to certain liquid components, analytes or interaction products of analyte, thereby permitt~ng only selected species to come into fluid contact with particular zones of a multizone element, such barrier compositions being described in US Patent 4,066,403; and zones including rupturable pod-like mem-bers which contain a liquid interactive composition as a reagent to be released in~o the elPment upon rupture .~
, .~ ..
~, , .
,, - .
. ' `~ ' .

of the pod-like member, such 70n2S being described in US Patent 4,110,079.
Methods of preparing and incorporating the above-noted zones in multizone elements of the inven-tion are id~ntical or similar to such methods as described in the foregolng US patents. Description of useful materials ~hich can be employed to prepare such optional zones or layers are also provided in the ore-going patents.
Typically~ except for reflecting and radiation-blocking agents, zones or layers which may be present in elements of the invention, the various zones, supports and other layers which may be present in an element of the invention are "radiation-transmissive". In the present specification, ~he term "radiation-transmissive" refers to zones, supports 9 layers and other materials in an element which permit effective passage of electromagnetic radiation used to detect an analytical change produced in the element.
Such radiation can include visible light, fluorescent emission, radioactive radiation, X-ray radiation and the like. The choice of a particular "radiation-transmissive" materlal in any given instance will depend upon the particular radiation selected for use with an element in which the material is to be incorpo-rated. Of course, radiation-transmissive materials are not required in the present invention. In various embodiments, one may choose to use radiation-blocking agents, zones and layers to prevent radiation from interfering with certain chemical interactions occur-ring within an element of the invention, e.g., interac-tions involving radiation-sensitive materials.
As noted above, the varlous zones or layers of an analytical element of the invention are in "fluid contact" with one another. In the present specifica-tion, the term "fluid contact" and similar terms refer '"
, ,. . :' :
:' :
.
' ' $

-41a-to zones or layers of an element associated with one another in a manner such that, under conditions of use, a fluid, whether liquid or gaseous, can pass in the element between these layers or zones. Such fluid contact therefore refers to the capability of the element to permit passage of at least some com-ponents of a fluid sample between zones or layers of the elemen~ which are said to be in "fluid con-tact". Such fluid-contact capabili~y is preferably uniorm along the contact interface between the fluid-contacting zones. Zones which are in fluid contact can be contiguous, but they also may be . ., ~
, ' .
,. .

_42-separated by intervening zones or layers. Such intervening zones however will also be in fluid contact in this case and will not prevent the passage of fluid between the fluid contacting layers or zones. In many embodiments, zones or layers in fluid contact are contiguous with one another or separated by a mutually contiguous intervening zone prior to application of a liquid sample to the element. Nevertheless, in some circumstances it may b~ desirable to use initially spaced-apart zones or layers within an element as illus-1- trated hereinabove in the element of Fig. 9. In such case, fluid contact between such spaced-apart zones is ach~eve~
substantially at the time of sample application, as by applying a compressive force to the element.
As previously mentioned~ the elements of the 1, invention can be self-supporting or carried on a support.
Useful support materials include a variety of polymeric materials such as cellulose acetate, poly(ethylene tere-phthalate), polycarbonates, and polyvinyl compounds such as polystyrenes, glass or metallic supports, paper supports, 2 and the like. A support of choice for any particular element will be compatlble with the intended mode of result detection. For example, for fluorimetric detection wherein fluorimetric emission within the element ls detected as the emission is transmitted from within the element through the 2~ support to an external detector, it ls desirable to employ as a support material a material which exhiblts a low degree of background fluorimetric emission. Thus, pre~erred supports include supports which are radiation-transmissive with respect to the particular radiation employed to provlde 3v detectable changes within the element. Thus, again, in the case of an element which provldes a fluorimetrically detect-able change, it is desirable to employ as a support, a material which transmits radiation at both the absorption and emission spectra o~ a ~luorescent material used for result detection. In certain cases, lt may also be deslrable to have a support that transmlts one or more narrow wave-length bands of radiation and is opaque to ad~acent wave-length bands of radiation. This may be accompllshed~ for example, by impregnating or coating the support with one or more colorants or other opacifying a~ents having suitable absorption characterist~cs. Typically, when an element does include a support, the reagent zone, the re~lecting or radiation-blocking zone, and the reglstration zone (lr any one or more of the foregoing zones are present in the element), will usually, but not necessarily, be interposed in the element between the support and the particulate structure-containing layer or zone of the invention which 10 often is the outermost layer or zone in the element. In general, the components of any particular layer or zone of an element of the invention, and the layer or zone con~igura-tion of the element, ~ill depend on the particular use for which that element is intended.
In preparing multi-zone elements of this inven-tion, the individual zones can be preformed and thereafter laminated prior to use or maintained as separate zones until brought into fluid contact when the element is placed in use. Zones preformed as separate members, if coatable, can 20 advantageously be coated from solution or dispersion on a surface from which the zone can be physically stripped when dried. However, a convenient procedure which can avoid problems of multiple stripping and lamination steps when contiguous zones are desired, is to coat an ~nitial zone on 25 a stripping surface or a support, as desired, and thereafter to coat successive zones directly on or beside those pre-viously coated. Such coating can be accomplished by hand, using a blade coating device or by machine using technlques such as "dip" or "bead" coating. For example, where the 30 multi-zone elements represent elements bearing superlmposed multiple layers, these multilayer elements can be coated using sequential coating techni~ues or using simultaneous multilayer coating methods and apparatus well known in the photographic art, such as, for example 9 the methods and 35 apparatus described in United States patents 2,761,417, 2,761,418, 2,761,419 and 2,761,791. Use of simultaneous multilayer methods of coating is often advantageous in that it avoids the problem of "air-cratering" which can arise _44-when layers are coated sequentially with a dr~lng s~ep between the coating of each successive layer. This problem results from the fact that as a layer is coated, the liquid medium in the coating composition enters the voids in the underlying layer and displaces air which ruptures the overlying layer and causes "pockmarks" or "craters" therein.
Simultaneous multilayer coating is also adYantageous in that it generally provides a substantial saving in the time and expense involved in the coatin~ operation as compared to sequential techniques.
Slide-extrusion hoppers of the type described in Vnited States patent 2,761,417 are often advantageous for simultaneous coating of a plurality of layers at least one of which is comprised of the organo-polymeric par~icles des-cribed herein. More particularly, a multilayer element canbe coated by directing a coating composition containing the organo-polymeric particles through an extrusion slot of a slide-extrusion hopper and simultaneously flowing a layer of a second coating composition, which, if desired, may also contain organo-polymeric particles, down a slide surface of the slide-extrusion hopper. Preferably, the coating compo-sition flowing through the extrusion slot is supplied at a volumetric flow rate that is substantially greater than the volumetric flow rate of the coating composltion flowing down the slide surface. Also, lt is desirable that the coating composition directed through the extrusion slot have a vis-cosity which is substantially higher than the viscosity of the coating composition flowing down the slide surface and a surface tension which is at least about as hlgh and, most 3 preferably, somewhat higher. Control of the coating para-meters of flow rate, viscosity and surface tension in this manner aids in promotlng the ~ormation of discrete layers that are free from interlayer mixing and in avoiding the ~ormation of repellency defects.
Elements of the present invention can be adapted for use not only in the field of clinical chemistry, but in chemical research and in chemical process control laboratories.
In addition, the particulate structure of the invention can ,: ,: ,.

, , , ~, , , , ~ , ~ , " ~, ~

;4~

be associated with other functional zones or layers outside the ~ield o~ analytical liquid analysis, e.g., layers or zones o~ photographic elements, to generally provide a resultant element having enhanced liquid transport capa-bilities. Analytical elements of the invention are wellsuited for use in clinical testing of body fluids, such as blood, blood serum and urine, because in this work a large number of repetitive tests are frequently conducted and test results are orten needed soon arter the sample ~s taken. In analyzin& blood with the analytical element of this inven-tion, the blood cells may first be separated ~rom the serum, by such means as centrifuging, and the serum applied to the element. ~owever, it is not necessary to make such separa-tion. Wnole blood can be applied directly to the element.

,, : ..

.

:. : : . :, The presence of blood cells on the element will not ln-terfere with spectrophotometric analysis ir it ls carried out by reflection techn~ques, with light being transmitted through the support and reflected from a radiation-blocking zone or other reflecting zone such that detecting radiation does not intercept the cells. Of course, if it is desired to directly observe the color o~ blood cells~ such as in a direct hemo~lobin analysis, no such rerlecting layer is necessary. A particu~arly significant advantage of the integral analytical elements described herein is their ability to be used to analyze either serum, plasma, or whole blood.
As can be appreciated~ a variety of different elements, depending on the analysis of choice, can be pre-pared in accordance with the present invention. Elementscan be configured in a variety of forms, including elongated tapes of any desired width, sheets or smaller chips.
The prepared elements are placed in use by apply-ing to the element a sample of liquid under analysis.
Typically, an element will be formed such that an applied sample will first contact a zone having the described particulate structure to spread and transport the sample wlthin the element, for example, to an ad~acent or under-lying reagent zone, if such a zone is present in the element.
Because analytical accuracy of the present elements is not substantially diminished even though some variation ln the volume of applied samples is encountered, sample application by hand or machine is acceptable. For reasons of convenience in detecting an analytical result, however, reasonable consistency in sample volume may be desirable.
In a typical analytical procedure using the present elements, which could be manual or automated, the element is taken from a supply roll, chip packet or other source and positioned to recelve a free drop, contact spot or other form of liquid sample, such as ~rom an appropriate dispenser.
After sample application, and desirably after the liquid.
sample has been taken up by the particulate structure, the element ls exposed to any conditioning, such as heating, . . .

, . ~ , .
, .

humidification or the like, that may be desirable to quicken or otherwise facilitate obtaining any test result. If an automated procedure is used, it can also be desirable to have the part~culate structure accomplish its liquid trans-port and spreading function within 20 to 30 seconds,preferably 20 seconds or less.
After the analytical result ls obtained as a detectable change, it is measured, usually by passing the element through a zone in which suitable apparatus for reflection, transmission or fluorescence spectrophotometry, or scintillation counting is provided. Such apparatus would serve to direct a beam of energy, such as light, through the support. The light would then be reflected, such as from a radiation-blocking layer in the element, back to a detecting means or would pass through the element to a detector, in the case of transmission detection. In a prererred mode, the analytical result is detected in a region of the element totally within the region in which such result is produced.
Use of reflection spectrophotometry can be advantageous in some situations as it can effectively avoid interference from residues which may have been left on or ln the layers Or the element. Conventional techniques of fluorescence spectrophotometry can also be employed if the detectable change produced in the element represents an increase or 25 decrease in fluorescence. Detection would be accomplished using energy that excites a fluor and a detector that senses its rluorescent emission. Furthermore, when blood serum is tested, transmission techniques can be used to detect and quantify the released indicating ligands by directing a ~low 30 of radiant energy, for example, visible radiation, at one surface of the element and measuring the output Or that energy from the opposing surface of the element. Generally, electromagnetic radiation in the range of from about 200 to about 900 nm has been found useful ~or such measurements 5 35 although any radiation to which the element is permeable and which is capable Or quantlfying the detectable change produced in the element can be used. Various calibratlon techniques can be used to provide a control for the analysis.

.

As one example, a sample of analyte standard solution can be applied ad;acent to the area where the drop of sample is placed in order to permit the use o~ dif~erentlal measure-ments in the analysis.
Immunoassay This Section discusses a specific application Or the particulate structure described herein, namely lmmuno-assay. This application represents an especially preferred embodiment of the invention.
Immunoassay is a well-recognized technique for qualitative or quantitative assay of antibodies and antigens.
Tne basis ror all immunoassay techniques is the unique, immunological phenomena whereby a speciric antibody recogniæes and binds to a specific antigen. Immunochemic~l techniques 1~ offer advantages in terms of assay sensitivity because of the high affinity of antibody for its specific antigen.
Therefore, in many instances immunoassay has made possible the detection of biological compounds that are present in trace quantities too low for traditional chemical and enzymatic techniques.
In general, immunoassay techniques can provide for a determination o~ the presence and/or concentration o~
either a specific antigen3 a specific antibody, or a specific antigen-antibody complex. For example, given a known amount o~ antibody (or antigen), the level of its corresponding antigen (or antibody), sometimes rererred to as its complement, can be determined. When the concentration Or antigen (or antibody) is too small for direct measurement, a label (i.e., detectable species) can be af~ixed to a known 3 fraction Or the antigen (or antibody). This label, which is present and measurable at the requisite concentrationl acts as a marker ~or the extent o~ antibody/antigen binding between the unknown antigen (or antibody) and its antibody (or antigen). The distribution of label between the bound 3~ and unbound antigen (or antibody) can then be ùsed ~o calculate the amount of unknown that was present in a liquid test sample.

..

To accomplish the foregoing determination, many current immunoassay techniques require the physi-cal separation of bound and unbound antigens (or anti-bodies); this is an additional step in analysis which can be inconvenient and time-consuming. Also, most currently available techniques suffer from one or more of the following disadvantages: (a~ relatively large volumes (~.1-1.0 ml3 of serum or other test liquid may be necessary compared with conventional chemical and enzymatic assays typically using 1.0-200 ~1 of liquid sample; (b) time-consuming incubation (several hours or overnight) of the test mixture is required; ~c) many steps are necessary and must be performed individually and separately for completion of the assay (including sample addition, incubation, separation, quantitation of label); (d) tests must o~ten be run batchwise; and (e) adaptation to an automated system is difficult.
Use of an analytical element with a particulate structure described above to conduct immunoassay o~er-comes many of the above drawbacks. The basic principles of specific binding of antigen to antibody are embodied in these immunoassay elements, the preferred immunoassay elements described hereinafter relying particularly on competitive binding of a labeled and unlabeled antigen (or antibody) to its specific antibody (or antigen). It will be understood, however, that an immunoassay element comprising a particulate structure described above can be made within the scope of the invention, relying on basic principles of antigen-antibody interaction other than competitive binding. For example, an immunoassay based on an antigen-antibody displacement interaction as described in US Patent 4,166,093 may be conducted with an immunoassay element comprising in one zone thereof a particulate structure and, in association with that zone, a labeled antigen-antibody (or antigen-labeled antibody) complex. The presence and/or concentration o~
an unknown antigen (or antibody) is determined by dis placement of the labeled antigen (or labeled antibody) . . . , -, . :
,''' ~ .

from the preformed labeled an~igen-antibody (or antigen-labeled antibody) complex.
For illustrative purposes and for purposes of describing the currently preferred mode oE immunoassay element comprising a particulate structure, the remain-der of this section is directed to an immunoassay ele-ment for ~he determination of the presence and/or con-centration of an antigen based on the competitive bind-ing of that unlabeled antigen and a labeled antigen to its antibody.
Thus, for example, a known amount of an antigen is rendered detectable, i.e., labeled, with a detectable species, such as with enzymes or fluorescent species or radioactive species. The antigen can be chemically linked or physically adsorbed to the detectable spe-cies. For example, a fluorescent species such as fluo-rescein can be covalently bonded to the antigen. In a preferred embodiment, a polymeric latex bead is "loaded"
with a rare-earth chelate~ a fluorescent species, and the resultant rare-earth-chelate-loaded latex bead is employed as a fluorescent label ~o which ~he antigen of choice is physically adsorbed or co~alently bonded.
These latex polymer beads typically have an average diameter of from about 0.01 to about 0.2 micron and are "loaded" with up to about 7.5 weight percent of a rare-earth chelate, preferably a europium or terbium che-late. Because of the large number o rare-earth chelate molecules which can be loaded into a single latex bead, the resultant label is highly fluorescent and provides a fluorescent immunoassay exhibiting excellent sensi-tivity. Labeled antigen employing a fluorescent, rare-earth-chelate-loaded polymeric latex bead as the label is described inCanadian Serial No 316,642.
Also, an amount of the antibody for the labeled antigen is incorporated and immobilized in an analytical element preferably within a zone thereof comprising a particulate structure. Such immobilization is accom-plished by adsorption or chemical bonding of the anti-body to the surface of the organopol~meric particles ', ` ~; ' , o~ the particulate structure. The liquid sample to be analyzed ~or unknown antigen is then contacted together with the element in the presence Or the labelled antigen. The labelled antigen may be associated with the immunoassay element in one of several ways including~ among others:
direct addition Or the labelled antigen to the liquid sample (containing unlabelled antigen) which is then applied to the i~munoassay element for analysis; separate addition of the labelled antigen and the liquid sample to the immunoassay element, including addition of the labelled antigen ~ust prior to or after addition of the liquid sample as well as addition of the labelled antigen to the element followed by drying and then rewetting the element upon addition of the liquid sample to be tested; or incorpora~ion o~ the labelled antigen in the immunoassay element so that analysis can be initiated simply by application of the liquid sample to be tested. For example, the labelled antigen may be incorporated in a separate reagent zone Or the element or in the same zone of the element containing the immobilized antibody. In any case, when the labelled antigen is incorporated in the element, care should be taken to maintain the labelled antigen apart from the immobilized antibody also in the element so that premature binding of labelled antigen to antibody is avoided.
When the liquid sample is contacted together with the immunoassay element in the presence Or the associated, labelled antigen as noted above, the labelled antigen and the unlabelled antigen (present in the sample and represent-ing the unknown to be determined) compete for binding to the 3 antibody which is present immobilized in one zone of the element. ~seful methods of measurement to determine the presence and/or concentration of unlabelled antigen which can then be employed include: (A) detecting the unbound, labelled antigen which has migrated into a second zone of 35 the element, e.g., Q registration zone9 or (B) detecting the bound, labelled antigen which blnds to the immobilized antibody. In either method, the amount of unlabelled antigen (i.e., the analyte) in the liquid sample can be determined :

based on the detected concentration of labelled antigen.
A partial listing of representative analytical elements illustrating various embodiments Or an immunoassay element containing a partlculate structure are presented hereinafter. Of course, as indicated above, other element configurations and other immunoassays may also be possible within the scope of the invention and therefore this listing is not exhaustive.
1. Fluorescence Immunoassay Element (FI~) This embodiment~ as shown in Fig. 9, comprises two superposed zones, each composed of a particulate structure of the invention, carried on a low fluorescence, radiation-transmissive support3 e.g., a ~lexible plastic support such as one composed of polycarbonate, cellulose acetate, or polystyrene. The particulate structure-containing z~ne immediately over the support represents a registration zone.
The particles of the registration zone are preferably spherical, organo-polymeric beads having a uniform size (i.e., they are monodisperse beads) within the range of from 5 to about 20 microns, most preferably about 6 to ~ microns~
in diameter. Typical organo-polymers for the bead composi-tions of this zone are polymers 2 and 6 of Table I, prefer-ably containing very low amounts of residual, unpolymerlzed styrene monomer, e.g., less than about 1% by weight (based 25 on the dry polymer) of residual styrene. A pre~erred adhesive for this particulate structure is organo-polymer 1 o~ Table II. Preferably, a nonspecific protein such as ovalbumin, bovine serum albumin, gelatin, diluted nonimmune serum, etc., is adsorbed to the particles of the registra-3 tion zone to minimize nonspecific binding in the finalassay. In addition~ this zone may optionally contain a highly reflective component, for example, from 1 to about 25 percent by weight of a pigment such as TiO2 or BaS04. This amount of pigment can enhance light scattering withln the 35 registration zone, thereby efrectively increaslng the light available within the zone to excite a fluorescent-labelled species which migrates into this zone for detection.
Viscosity modifying agents and surfactants may also be ,~ :
. .

contained in this zone to ~acilitate its preparation as described hereinabove. A bu~fer can also be employed ln this zone to maintain its pH, under conditions Or use, between about 7 and 9. The upper zone represents a spread-ing/reagent zone and the particles in this zone also have a uniform size, pre~erably similar to that Or the particles in the re~istration zone, and have antibody immobilized thereon, e.g., by adsorption or chemical bonding. Thls spreading/re-agent zone may also contain a nonspecific protein as con-lc~tained in the registration æone. The upper zone providesfor unifor~ spreading of an applied liquid test sample, and the high surrace-to-volume ratio of the particulate struc-ture ~orming this zone gives excellent binding capacity. A
pigment or dye can also be incorporated in some or all of 15 the particles o~ this zone to serve as a radiation-blocking agent, i.e., a light screen, for a fluorescent species. A
partial listing of representative such dyes or pigments includes Wachtung Red B Pigment~ (from E.I. duPont deNemours), Permanent Purple~ (~rom GAF), Sol Fast Methyl Violet~ (~rom 20 Sherwin-Williams), Indofast Blue~ (from Harmon Colors), Regal 300~ (from Cabot), Monolite Blue~ (~rom ICI), and Paliofast Blue~ (from BASF). The inclusion of a viscosity modifying agent and surfactant is optional. A buffer to maintain the spreading/reagent zone under conditions o~ use 2~ at a pH between 7 to about 10, pre~erably about 8.5, is typically present also. To carry out an immunoassay with this element, a ~luorescent-labelled antigen may be associ-~ted with the element as mentioned earlier in this Section.
For example, the labelled antigen may be lncorporated in an 3 optional reagent zone together with any necessary or deslre-able binder or applied to the element together with the liquid test sample. A~ter applying the liquid test sample to the element ln the presence o~ the assoclated, labelled antigen, the sample contac~s the reagent/spreading zone o~
the element and the competltive binding interaction among the fluorescent-labelled antigen~ the unlabelled antigen in the liquid test sample representing the analyte, and the immobili2ed antlbody takes place. The unbound ~luorescent-labelled antigen migrates into the registration ~one where ., .

6 ~ ~
-54_ it can be quantitated through the clear plastic support by illuminating the zone with li~ht at the excitatlon ~avelength of the ~luorescent label and measuring the emitted ~luores-cence. The radiation-blocking agent in the spreading/re-agent zone hides the fluorescent label that remains bound tothe immobilized antigen-antibody complex remaining thereinO
2. Monolayer Fluorescence Immunoassay Element As shown in ~ig. 11, another type Or immunoassay element containing a particulate structure comprises a 10 monolayer Or antibody directly adsorbed to a plastic support similar to that used in FIA embodiment tl) above. This ca~
be accomplished by incubating the plastic support with a solution Or antibody (e.g., antiserum) diluted by a ~actor of from about 10 to 5000 (depending on the concentration of 15 antibody in the antiserum and the range of the assay~ for a period o~ several minutes to several hours, e.g., 0.5 minutes to 48 hours. The support can then be rinsed in saline and incubated in a nonspeci~ic protein solution to minimize nonspecific binding in the assay. This second 20 incubation step can be carried out in a time period similar to the first incubation step. The resultant support can then be rinsed in water and air-dried. The spreading zone is the same as the spreading/reagent zone ror embodiment (1), except that no antibody is adsorbed in this zone.
25 Labelled antigen may be associated with the element as in embodiment (1) above. In operation, the amount of labelled antigen bound to the adsorbed antibody is quantitated; the radiation-blocking agent in the upper spreading/reagent zone hides the unbound labelled antigen.
3. Radioimmunoassay Element (RIA) In the element shown in Fig. 12, a radiation-transmissive plastic support bears a scintillation zone as a registration zone which may be, for example, a solid late~
scintillator layer as described in Chen, Miller and Perry, 35 U.S. Patent No. 4,127,499, issued November 28, 1978, or, pref-- erably, a particulate structure o~ the present invention comprising particles bonded with a fluor-imbibed latex ad-hesive. Illustrative rluor-imbibed latex adhesives include . - : , . , the scintillation rluors 2,5-diphenyloxazole and 2,2'-p-phenylenebis(5-phenyloxazole) imbibed lnto a latex ~orm of one of the polymeric adhesives noted in Table II. Other components such as surfactantsg viscosity modifying agents, buffers and the like as noted above may also be used in preparing this particulate structure. Over the scintil-lation zone is a spreading/reagent zone containing a partl-culate structure with antibody adsorbed to the particles thereor as in the FIA element Or embodiment (1) above, except that no plgment is incorporated. The principles embodied in use Or this immunoassay element are the same as in the FIA element Or embodiment (1) above, except that the label is radioactive, rather than ~luorescent. The unbound radioactive-labelled antigen is detected in the lower 15 registration zone which is the solid equivalent of a "scintillation cocktail." Label in the upper spread-ing/reagent zone is not quantitated. This embodiment is particularly use~ul for the determination of very high molecular weight antigens because o~ its enhanced perme-20 ability and high counting erriciency.
4. Enzyme-Enhanced Immunoas:a_ Element In this embodiment as shown in Fig~ 13 above, the label is an enzyme. The radiation-transmissive plastic support is coated wlth a reagent zone containing portions o~
25 an interactive enzyme assay composition for the ~luoro-metric, colorimetric or radioactive detection Or an enzyme.
This enzyme assay composition may be admixed ln a film-forming vehicle such as gelatin, hardened gelatin, or agarose. For example, if the enzyme label is peroxidase, 3 the enzyme assay composition may contain glucose, glucose oxidase (to generate peroxide from glucose) and a reduced dye precursor which is oxidized in the presence of peroxide and peroxidase, thereby producing a radiometrically detect-able change in ~he element. The glucose portlon o~ this 35 enzyme assay composition can be located in a zone separate from the enzyme assay-containing reagent zone, for example, the spreading/reagent zone of Fig. 13, to avoid premature reaction of glucose with glucose oxidase. Or, the glucose portion could be separately added to the element or added . ~

to~ether with the liquid test sample. At the time of use, the liquid test sample applied to the element causes the glucose to migra~e into the enzyme assay-containing reagent zone to interact with the other components of ~he enzyme assay composition. Various surfactants can also be ln corporated in this reagent zone compositlon to ald coat-ability. The optional scavenger zone of Fig. 13 prevents enzyme substrate, e.g. H2023 generated in the enzyme assay-containing reagent zone from migrating up into the spreading~reagent zone where the enzyme labelled antigen, e.g., a peroxidase-labelled antigen, is present. As a result, only the unbound enzyme-labelled antigen that migrates into the enzyme assay-containing reagent zone is quantitated. Thus~ where the enzyme label is peroxidase, the scavenger zone may contain a film-forming binder (such as gelatin), buffer, and catalase (catalase decomposes H202) to prevent migration of H202 generated in the enzyme assay-containing reagent zone into the spreading/reagent zone where it could react with peroxidase-labelled antigen that is bound to its immobilized antibody. The spreading/reagent zone is identical to that in the RIA element of embodiment (3) above, except that a portion of the interactive enzyme assay composition employed in the enzyme assay-containing reagent zone of the element may also be present in thls zone, e.g., glucose as described immediately above. The . enzyme-labelled antigen may be associated with the element by any of the previously mentioned techniques.
5. Fluorescence Immunoassay Element for Low-Molecular-Weight Antigens 3 This modification of the FIA element of embodiment (1) above is illustrated in Fig. 14. The support ls the same as that in embodiment (1). The registration zone is composed of a polymeric vehicle such as gelatin or agarose, and coating agents such as surfactants. Bufrers or other components which might serve to enhance the ~luorescence of the labeiled antigen may be lncluded in the registration zone. The radiation-blocking zone is similar in compositlon to the registration zone but contains a dye or pigment as a radiation-blocking agent to screen out the ~luorescence Or .: ~, ~ ' ' ., ' labelled antigen bound in the upper spreading/reagent zone.
The spreading/reagent zone is the same as described for the FIA eleme~t of embodiment (1~, except that the particles in the part~culate structure of this zone may or may not be pigmented.
The following examples are presented to ~urther illustrate certain embodiments of the present invention. In each of these examples numbers appearing in parentheses following a polymer name, e.g., (~8:2~, represent the parts by weight of the respective monomer components contained in the initial monomer blend from which the polymer was pre-pared.
Example 1 Organo-Polymeric Bead Structure For Transport Of Whole Blood In this part of Example 1, three particulate aqueous transport layer structures were prepared and tested for whole blood transport capability. Two of these layer structures were control structures of the type specifically described in U.S. Patent 3,992,158 and are outside the scope of the present invention. The third structure was an organo-polymeric particulate structure of the present invention. The two control layer structures were chosen on the basis of their demonstrated excellent performance as aqueous transport structures ror blood serum. The first control layer structure tested was composed of a "blushed"
cellulose acetate polymer layer containing TiO2 particles.
This particulate layer structure had a composition and was prepared in a manner similar to that described in detail in Example 3 of U.S. Patent 3,992,158. The second control 3 layer structure was composed of Avicel particles (micro-crystalline cellulose particles purchased from FMC Corp.) dispersed in a binder polymer consisting of poly(vinyl pyrrolidone). This layer structure was coated from an aqueous coating mixture. The amount of microcrystalline cellulose particles contained in this layer structure ~based on dry weight) was about 64.5 g/m2 and the amount of poly~vinyl pyrrolidone~ was about 1.~ g/m2 talso based on dry weight). The approximate size of the microcrystalline cellulose particles in the second control structure was , . , ~
: . , ;
- -about 30 to 50 microns.
The third structure evaluated in this example was an organo-polymeric particulate layer of the present lnven-tion coated from a water dispersion having the following dry composition:
Coati~g Covera~e (i) 97.8 parts by weight Or solid, spherical organo-polymeric beads o~ poly(styrene-co-di-vinylbenzene) (98:2) having a bead size of from about 35 to 75 microns 196 g/m (ii) 2 parts by weight of a polymer adhesive composed of poly(n-butyl acrylate-co-styrene-co-2-acrylamido-2-methylpropane sul~onic acid) (70:20:10~ 4 g/m (iii) 0.2 part by weight of Surfactant lOG (p-nonyl phenoxy polyglycidol purchased from Olin Corp.) o,4 g/m2 The polymer adhesive employed in the above-described composi-tion was prepared as an aqueous latex having a 33.2 weight percent solids content consisting of the above-mentioned copolymer as a discontinuous phase and water as the con-3 tinuous phase. This latex was pre-formed and then diluted 16.5X with water. A water dispersion containing the organic polymeric beads (i) and Surractant lOG (iii) was then added to the adhesive latex and the resulting water dispersion was used to coat the above-described organo-polymeric bead 35 structure. The wet coating thickness of the structure was about 530 microns. Subsequent to coating, the bead structure was air-dried at 54C. The dry thickness of the resultant bead layer structure was about 370 microns.

' . ' : , : . , . , , -.

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Each of the three above-described layer structures was coated on a plastic film support of poly(ethylene terephthalate) bearing a thin film subbing layer to aid adhesion. The structures were then evaluated for whole blood transport capability by applying constant drop-size samples ( ~10 microliter drops) of whole blood to each structure and analyzing the ability of each structure to take upg uniformly distribute within itself, and rapidly transport the whole blood drop samples.
The results of this evaluation demonstrated that the organo-polymeric particulate structure of the present invention provided uniform and rapid transport of the blood drop samples. The red blood cells in these whole blood drop samples were rapidly taken up and uniformly distributed 15 within the layer in approximately 10-20 seconds and the structure became uniformly pigmented. Moreover, spot size of the whole blood drop samples on the bead structure of the invention was uniform and reproducible, spot sizes of about 10 to 12 mm being obtained. Microscopic examination of this 20 layer structure revealed an approximate mean void size of 25 microns.
In contrast, the control aqueous transport struc-tures demonstrated substantially less effective whole blood transport capability. Typically a portion of the red blood 25 cells were not taken up by control structures but were retained on the surface of the structure while the plasma was absorbed into the interior of the structure, or the blood cells were lysed by the structure and the resulting products of the lysis were taken up by the structure in an 3 irregular manner as demonstrated by the non-uniform spot sizes and shapes produced by the whole blood drop samples on these control structures.
The results of this example demonstrate that the organo-polymeric particulate structures of the present 35 invention are able to effectively and conveniently transport and accommodate aqueous,samples containing such large and complex physiological species as red blood cells.

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-6~-Example 2 Organo-Polym ric Bead Structure for Whole Blood An organo-polymeric particulate structure of the present invention was prepared in a manner similar to that described for the third layer structure in Exam-ple 1 above except that the solid, spherical beads were about 65 to 120 microns in diameter and were composed oE
poly(styrene-co-methacrylic acid~ (98:2), and the struc-ture was coated out of an aqueous dispersion. This bead s~ructure was evaluated as in Example 1 for transport of whole blood and was also found to provide an extremely effective whole-blood transport structure.
The organo-polymeric beads o~ this Example were prepared by bead polymerization as follows:
Bead Polymerica~ion of Styrene and Methacrylic Acid A Materials:
Styrene, methacrylic acid, potassium dichromate and 2,2'-azobis(2-methylpropionitrile) (AIBN) were used. In addition, colloidal silica was used as obtained from duPont either as a 3~ or 40% solution with the trade name Ludox HS30'~ or Ludox HS4DT~. A surface-active agent was also used consisting of a condensation copoly-mer of adipic acid and diethanol amine prepared by heat-ing the monomers neat, in a beaker equipped with a mag-netic stirring bar, on a hot plate until a centipoise reading of 12,000 to 350,000 was obtained using a Brook-field viscometer, Model LVT, 24 C, 3-0.6 rpm, and a number 4 spindle.
B Procedure:
-1 Aqueous Phase: Water (600 g), Ludox HS3~ (90g), diethanolamineadipic acid copolymer (15 g of a 10%
aqueous solution) and potassium dichromate ~6 g of a 2.5% aqueous solution~ were placed in a 2000-ml brown bottle, and the pH was adjusted to 4 with a l N
hydrochloric-acid solution.
2 Organic Phase: Styrene (588 g), methacrylic acid (12 g) and AIBN (6 g) were placed in a -Elask and stirred until the intiator had dissolved.
_ Dispersion: The organic phase was dispersed in the "

aqueous phase (with cooling) by a Brinkman Polytron homogeni-zer (purchased from Brinkman Instruments Company); 11~
volts, 5 amps, and 60 Hz, ror 2 minutes at a setting of 5.
Auxll~ary stirring with a Lightnin mixer (purchased ~rom Lightnin Company) was required to ob~ain a uniform dls-persion. The brown bottle was capped, sealed with tape, allowed to remain at ambient temperature without agitation ~or one hour to effect limited coalescence, and finally placed in a 60C bath overnight.
The following day the brown bottle was removed from the bath, allowed to cool, the reaction mixture stirred to redisperse settled beads, strained through 2 rine mesh screen, and collected on 230 Reeve Angel filter paper (purchased from Reeve Angel Company). The beads were 15 redispersed in water three times, collected after each redispersal, and air dried a~ter the final redispersal. The dried beads were put through a 100 mesh sieve to remove any large particles that were present.
Example 3 Multi-Zone Organo-Polymeric Bead Structures Fôr Whole Blood Transport And Red Cell Separation -In this example, a multi-zone element of the type illustrated in Fig. 4 is demonstrated as follows:
A poly(ethylene terephthalate) ~ilm support bearing a thin adhesive subbing layer was overcoated wlth two, superposed 25 organo-polymeric bead structures (i.e. layers) of the present invention. The structures dirfered with respect to the si2e of the beads used in the structures. The bead structure immediately ad~acent the subbing layer of the ~ilm support consisted of a bead structure slmilar to that described in 3 Example 2, except that the poly(styrene-co-methacryllc acid) beads were about 6 microns in diameter. A second bead structure was then coated over this rlrst bead structure.
The second bead structure was ldentical to that described in Example 2. The multi-zone element of this example was the~
3~ tested for its blood transport capabllity. It was ~ound that the top layer containing the large beads readily took up and uni~ormly distributed the whole blood drop samples applied to it as evidenced by the uniform coloration formed ~ . .

:

.

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in the top layer a~ the site of contact with the whole-blood drop sample. The lower bead layer (containing the 6-micron beads) rapidly became wetted with colorless por-tions of the whole-blood sample but did not take up any of the red blood cells as evidenced by the ~act that it was a clear layer and remained so, even after evaporation of the fluid portion of the whole-blood drop samples. This exam-ple demonstrates the capability of organo-polymeric bead structures to be used in a multizone analytical element to separate components of an aqueous liquid into two or more distinct zones based on the molecular size and configura-tion of the components.
Example 4: Multizone Element for Determination of Whole Blood or Plasma Glucose Using Organo-Polymeric Bead-Transport Structure In this example, a multizone element for determi-nation of whole blood or plasma glucose was prepared. The element was a multizone element of the type illustrated in Fig 7. The element had a transparent poly(e~hylene tere-phthalate) film support bearing a thin adhesive subblng layer overcoated with the following layers (listed in order beginning with the layer immediately adjacent the aforementioned subbing layer):
(i) Color-forming en2ymatic glucose reagent layer containing glucose oxidase (24,000 U/m2), 4-amino-antipyrene hydrochloride (0.86 g/m ), 1,7-dihydroxy-naphthalene (0.65 g/m2), peroxidase (18,000 U/m2), 5,5-dimethyl-1,3-cyclohexanedione (0.22 g/m2), 6-amino-4,5-dihydroxy-2-methylpyrimidine (0.02 g/m2), deionized gelatin (16 g/m2) and 3,3-dimethylglutaric acid (1.96 g/m2) as buffer to maintain pH of reagent layer at 5.0 when the element is spotted with drop of whole blood;
(ii) Subbing layer containing poly-n-isopropylacryl-amide (0.32 g/m );
(iii) Radiation-blocking reflecting layer containing TiO2 (18 g/m2); Triton X-lOO~N ~ an octylphenoxy polyethoxy ethanol available from Rohn and Haas Co (1.08 g/m2); and poly(acrylamide-co-ethyl acryloyl-acetate (90:10) (10.8 g/m2 ); and :
`
.
. -`

(iv) Organo-polymeric bead structure to spread whole blood sample having a layer structure identical to that described in Example 2.
Calibration curves for the above-described multl-zone element were then generated by spottlng commerciallyavailable serum calibrator solutions containing rrom 0 to 400 mg glucose/dl. on the element, incubatlng the element for 12.5 minutes at 37C., and thereafter detecting the color density produced in the color-~orming reagent layer by 10 reI~lection spectrophotometry through the transparent poly-(ethylene terephthalate) film support. Two serum control ~luids and a whole blood sample, each containing an unknown glucose level, were then spotted on separate but identical samples Or the element and the glucose levels were deter-15 mined on each of these element samples by use of the pre-viously generated calibration curves. The glucose levels Or these unknowns were also determined by accepted reference methods. Good agreement was obtained between the glucose values predicted by the multi-zone element and those ob-20 tained by the reference method as shown in Table III below:

TABLE III
.
Reference Glucose Level Glucose Predicted By Level Multi-zone Sample Tested (mg/dl) Element (mg/dl) Serum Control Fluid (1) 89.8 94 Serum Control Fluid (2) 276 267 Whole Blood Sample ~6 93 Example 5 Organo-Polymeric Bead Structure As A Trans-port And Rea~ent-Containing Structure In this example, an element for the analysis of serum glucose was prepared. The element contained a single 35 organo-polymeric bead structure whlch also included an - interacti~e composition containing all the necessary color forming and enzymatlc reagents to perform a quantitative serum glucose assay. The bead structure was carrled on a ., ... .

poly(ethylene terephthalate) film support bearing a thin adheslve subbing layer. The element was prepared as follows:
Materials For Organo-Polymeric Bead Structure (i) Beads - Poly(styrene-co-vinylbenzyl chloride-co-methacrylic acid) (78:20:2);
(ii) Adhesive - Poly(n-butyl acrylate-co styrene-co-2-acrylamido-2-methylpropane sulfonic acid) (76:19:5);
(iii) Glucose oxidase - 206 units/mg. Lot No. 58 (Miles Laboratories), (iv) Peroxidase - 1000 units/mg (horseradish) obtained from Miles Laboratories (Cat. No. 38-444);
(v) Triton X-100 - Octylphenoxypolyethoxy ethanol obtained from Rohm and Haas Co. and (vi) Chromogen - 2-(3-bromo-5-methoxy-4-hydroxy-phenyl)-4,5-bis(4-methoxyphenyl)imidazole.
Preparation Of Bead-Enyme Slurry 20 g of beads were added to 100 ml phosphate buffer solution, pH 7.4. 5 ml of glucose oxidase tcon-taining 30 mg of protein) and 30 mg (30,000 units) of peroxidase were added. The mixture was stirred overnight at room temperature and washed on a filter funnel with 1000 ml of 0.15 N sodium chloride.
Preparation Of Element The subbed poly(ethylene terephthalate) support 25 was coated with the bead slurry described above (162 g/m2) 3 adhesive ~3.24 g/m added as a 33.2% aqueous dialyzed latex), Triton X-100 (0.4 g/m2) and chromogen (1.6 g/m2). The element was buffered to pH 5.9 with NaH2PO4.
The element was tested, qualitatively, by spotting 30 10 microliters of serum ~lucose standards containing 0, 15, 30, 60 and 125 mg/dl respectively, and Interstate Blood Bank serum calibrators (levels 1, 3, 7 and 8) and incubating at 37C. for 10 minutes. Results demonstrated good differ-entiation between each of the levels of glucose tested.
35 Example 6 Organo-Polymeric Bead Structure For Direct Hemoglobin Analysis _ _ It is known that the blood protein hemo-globin (Hb) can be assayed directly by analyzing spectro-s~

photometrically a dried sample Or whole blood spotted onto an absorbent matrix. Such a direct Hb assay is typically carried out by evaluating the 540 nm absorption peak oP Hb.
Although it is known that several different forms of hemo-globin exist, such as carboxy-hemoglobin, met-hemoglobln~ or sulf-hemoglobin, which do not exhibit an absorption peak at 540 nm, only minor amounts of such hemoglobin derivatives are generally present in whole blood. For example, the total Hb content of whole blood typically is composed of over 95 weight percent of oxy-hemoglobin having an absorp-tion peak at 540 nm and less than 5 weight percent Or other forms of Hb having absorption peaks at points in the visible spectrum other than 540 nm. Thus, an accurate total Hb assay can be performed merely by examining the 540 nm absorption band of a whole blood sample and ad~usting the result obtained by a standardized quantity to allow for the presence of minor amounts of other forms of Hb. Of course, this can be effective only if one has a suitable absorbent matrix on which to spot whole blood samples. Such a matrix 20 should rapidly take up a sample drop of whole blood applied thereto, hemolyze the red blood cells contained in the blood so that the Hb content of the cells is released, and uni-formly distribute the products of the lysis throughout the matrix in the form of a spot pattern exhibiting a regular 25 and reproducible shape and size. The dried spot pattern should show little or no irregularities and nonlinearities in color distribution across its surface area. Preferably, the absorbent matrix should perform the complete operation of whole blood hemolysis and spreading within itself in a 3 matter of seconds so that the Hb assay can readily be performed without delay. In this regard the organo-polymeric bead structures of the present invention have been found to represent an excellent absorbent matrix for per-forming Hb assays. A typical bead structure of the inven-35 tion for such a direct Hb analysis had a composition and wasprepared in a manner identical to that described in Example 2, except that Surfactant lOG was replaced by an amount of Triton X-100 ad~usted to provide 1 wei~ht percent of Triton : '., :

X-lOQ in the bead structure (as measured on a dry weight basis) and the amount Or the polymer glue contained in the bead structure was adjusted to 3 weight percent (also as measured on a dry weight basis). Triton X-100 was used to enhance hemolysis of the red blood cells. The "spread time"
required for this bead structure, i.e., the time required for it to take up, lyse the red blood cells and achieve uniform distribution of the lysis products throughout the structure was about 18 seconds. Even faster spread times of about 7-9 seconds were obtained using an organo-polymeric bead structure similar to that described above, except that the beads were prepared from a copolymer of poly(styrene-co-divinylben~ene) (98:2) as described in Example 1 having a bead diameter of from about 35 to 75 microns.
Examples 7-49 Alternative Organo-Polymeric Bead Structures A series of effective liquid transport elements were prepared in a manner similar to that described in Example 1, except that the organo-polymeric beads, the adhesive, and surfactant were varied as indicated in Table IV below. Elements 8-36 and 46 o~ Table IV are examples of elements as illustrated in Fig. 3 having a single layer of particulate structure carried on a support.
Elements 37-45 and 47-49 of Table IV are examples of elements as illustrated in Fig. 4 having a first particu-late structure layer 3 on a support 2 overcoated with asecond particulate structure layer 4.

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Example 50 Fluorescence Immunoassay Element A. Element Preparati In this example a fluorescence immunoassay element representing embodimen~ 1 as described in the Immunoassay Section was prepared.
1. Spreading/Reagent Zone Dispersion: An aqueous dispersion was prepared containing 46% (wt. %) microbeads adsorbed with the antibody antibovine gamma globulin (Anti-BCG); 0.5% (wt. %) of the non-ionic surfactant Zonyl FSN
(purchased from duPont)5 2.5% (wt. %~ polymeric adhesive No.
3 of Table II: and 2% (wt. %) normal rabbit serum as non-immune serum. The dispersion was buffered to pH 8.5 with a mixture of H3B03 and KCl. The viscosity of the dispersion, as determined by a Brookfield viscometer at 60, 30 and 12 rpm and room temperature, was 33 CP, and the surface tension of the dispersion was 24.9 dynes/cm2. The microbeads used in the aforementioned dispersion contained encapsulated Paliofast Blue as a radiation-blocking agent, had an average particle size of 6 to 8 microns, and were composed of poly-mer No. 2 of Table I. The amount of Paliofast Blue pigmentencapsulated by the microbeads was about 1.5 wt. % of the beads. Pigment encapsulation was achieved by incorporating the pigment into the Organic Phase of the bead polymerization described above in Example 2. Antl-BGG was adsorbed to the above-described microbeads by the following procedure: The beads were washed with 0.15 M NaCl to thoroughly clean the bead surface and then dried by suction filtration and resuspended at 20 wt. % solids in a solution containing 99 parts by volume of 0.03 M Na2C03~ pH 9.5, and 1 part by 3 volume o~ Anti-BGG rabbit serum. The bead suspension was stirred at room temperature for 24 hours and centrifuged for 10 min. at 6000 rpm to collect the beads. The beads were washed with 0.15 M NaCl and then centrifuged again.
2. Registration Zone Dispersion_ An aqueous dispersion was prepared containing 46%
(wt. %) microbeads adsorbed with normal rabblt serum; the microbeads having an average particle size of 6 to 8 microns and composed of polymer No. 1 of Table I; 2.5~ Cwt. ~ of polymeric adhesive No. 3 of Table II; and 0.2% (wt. ~ of the non-ionic surfactant Zonyl FSN . The dispersion was buffered as in (1) above. The viscosity o~ the dlspersion was 22 CP, and the surface tension was 29.5 dynes/cm .
3. Coating Procedure The immunoassay element was then prepared by coating as follows: A transparent polystyrene plastic film support exhibiting a low level of fluorescence was coated 10 with 80 g/m2 of the registration zone dispersion and 28 g/m2 of the spreading/reagent zone dispersion using the multiple slide-hopper bead coating technique described above in the Element Structure Section.
B. Analysis A 10 ~1 droplet of aqueous test solution buffered to a pH of 7.4 and containing 5 x 10 8 M fluorescein-labelled bovine gamma globulin as the labelled antigen and a varying level of unlabelled bovine gamma globulin as the unlabelled antigen ranging ~rom 0 to 10 5 M was applied to a 20 series of the immunoassay elements described in Part A. The buffer contained in the aqueous test solution was composed of 50% normal rabbit serum and 50% phosphate buffered saline (0.15 N saline, 0.01 M sodium phosphate). The 10 ~1 droplet of test solution was readlly taken up, i.e., spread~ by the 25 spreading/reagent zone in about 25 seconds. Thereafter the element was incubated for 15 minutes at 37C. A reflectance fluorimeter having excitation and emission filters at 490 and 515, respectively, was then used to obtain the data shown in Table V. The fluorimeter was set up to direct a 30 beam of light of the exci~ation wavelength of 490 nm through the polystyrene support of the element into the reglstration zone to detect fluorescence produced in this zone by the unbound labelled bovine gamma globulin which had mlgrated into the zone. ~hese fluorescence levels were then corre-35 lated to the varying levels of unlabelled antigen which wereknown to be present in the 10 ~1 of aqueous test droplets.
As can be seen from the response data in Table ~, these immunoassay elements produced a readily detectable change in fluorescence response corresponding to the varylng levels Or unlabelled antigen contained in the aqueous test droplets.
TABLE V
CONCENTRATION MEASURED
UNLABELLED FLUORESCENCE
BOVINE GAMMA (ARBITRARY UNITS~
GLOBULIN

2.5 x 10 8M 388 5 x 10 8M 431 1 x 10 7M 466 2 x 10 7M 491 1 1o~6M 54 1 x 10 5M 553 Buffer Blank 40 The invention has been described in detail with particular reference to certain preferred embodiments thereof, but it will be understood that variations and 20 modifications can be effected within the spirit and scope of the invention.

. . . , , . . .. ....... .: . .
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Claims (93)

We Claim:

1. A particulate structure for the analysis or transport of a liquid comprising:
(i) a plurality of heat-stable, organo-polymeric particles non-swellable in and impermeable to said liquid, said particles having a particle size of from about 1 to 200 microns, and (ii) an adhesive in an amount less than 10 percent by weight of said particles, comprising an organic polymer different from that of said particles and insoluble in said liquid;
said adhesive being concentrated at contiguous surface areas of adjacent particles and bonding said particles into a coherent, three-dimensional lattice which is non-swellable in said liquid and which has interconnected void spaces among said particles to provide transport of said liquid, said lattice having a void volume of from about 25 to 80 percent.

2. A particulate structure as defined in claim 1 wherein said organic polymer of said adhesive has a glass transition temperature less than that of said particles.

3. A particulate structure for the analysis of a liquid as defined in claim 1 wherein said particulate structure contains one or more components of an interactive composition for said analysis.

4. A particulate structure for the analysis of a liquid as defined in claim 1 wherein said particulate structure contains an immunoreagent.

5. A particulate structure for the analysis of a liquid as defined in claim 1 wherein the organo-polymeric composition of said particles comprises an organic polymer containing a repeating unit having a chemical group repre-senting an active bonding site for chemical attachment to an interactive composition for said analysis.

6. A particulate structure as defined in claim 1 wherein said particles are solid particles of substantially uniform size.

7. A particulate structure as defined in claim 1 wherein said particles are solid spherical beads having a substantially uniform size within the range of from about 2 to 100 microns.

8. A particulate structure as defined in claim 1 wherein the organic polymer of said particles and said adhesive comprise an addition copolymer of two or more different addition polymerizable monomers, at least one of said addition polymerizable monomers of said particles being common to at least one of said addition polymerizable monomers of said adhesive.

9. A particulate structure as defined in claim 1 wherein said particles contain a colorant.

10. A particulate structure for the analysis or transport of a liquid comprising:
(i) a plurality of heat-stable, organo-polymeric particles non-swellable in and impermeable to said liquid, said particles having a particle size of from about 1 to 200 microns, and (ii) an adhesive, in an amount of from about 1 to less than 10 percent by weight of said particles, comprising an organic polymer different from that of said particles and insoluble in said liquid;
said adhesive being concentrated at contiguous surface areas of adjacent particles and bonding said particles into a coherent, three-dimensional lattice which is non-swellable in said liquid and which has interconnected void spaces among said particles to provide transport of said liquid, said lattice having a void volume of from about 25 to 80 percent; said particles comprising an addition polymer of an addition polymerizable blend of monomers the total monomer content of said blend comprising at least one of the follow-ing monomers in the proportions indicated:
(a) from 0 to 100 weight percent of a polymerizable, amino-substituent-free styrene monomer;
(b) from 0 to about 25 weight percent of a polymeriz-able acrylic ester;
(c) from 0 to 100 weight percent of a polymerizable methacrylic ester;

(d) from 0 to about 30 weight percent of a carboxylic acid containing one or more polymerizable ethylenically unsaturated groups;
(e) from 0 to about 75 weight percent of a nitrile containing one or more polymerizable ethylenically unsaturated groups;
(f) from 0 to about 20 weight percent of a polymeriz-able amine-substituted styrene monomer;
(g) from 0 to about 20 weight percent of an addition polymerizable monomer containing a crosslinkable group;
(h) from 0 to about 20 weight percent of a polymer-izable tertiary aminoalkyl acrylate or methacry late;
(i) from 0 to 100 weight percent of a polymerizable, N-heterocyclic vinyl monomer;
(j) from 0 to about 20 weight percent of a polymer-izable acrylamide or methacrylamide monomer; and (k) from 0 to about 20 weight percent of a cross-linking monomer containing at least two addition polymerizable groups.

11. An element for the analysis or transport of a liquid, said element comprising a radiation-transmissive support bearing a particulate structure comprising:
(i) a plurality of heat-stable, organo-polymeric particles non-swellable in and impermeable to said liquid, said particles having a particle size of from about 1 to 200 microns, and (ii) an adhesive, in an amount less than 10 percent by weight of said particles, comprising an organic polymer different from that of said particles and insoluble in said liquid;
said adhesive being concentrated at contiguous surface areas of adjacent particles and bonding sald particles into a coherent, three-dimensional lattice whlch ls non-swellable in said liquid and which has interconnected void spaces among said partlcles to provide transport of said liquid, said lattice having a void volume of from about 25 to 80 percent.

12. A multi-zone element for the analysis or transport of a liquid, said element comprising, in fluid contact, a zone having a particulate structure and at least one other zone permeable to said liquid, said particulate structure comprising:
(i) a plurality of heat-stable, organo-polymeric particles non-swellable in and impermeable to said liquid, said particles having a particle size of from about 1 to 200 microns, and (ii) an adhesive, in an amount less than 10 percent by weight of said particles, comprising an organic polymer different from that of said particles and insoluble in said liquid;
said adhesive being concentrated at contiguous surface areas of adjacent particles and bonding said particles into a coherent, three-dimensional lattice which is non-swellable in said liquid and which has interconnected void spaces among said particles to provide transport of said liquid, said lattice having a void volume of from about 25 to 80 percent.

13. A multi-zone element for the analysis of a liquid as defined in claim 12 wherein one or more components of an interactive composition for said analysis is located in at least one of said zones.

14. A multi-zone element for the analysis of a liquid as defined in claim 12 wherein at least one of said zones contains an immunoreagent.

15. A multi-zone element as defined in claim 12 wherein the organic polymer of the adhesive in said particu-late structure has a glass transition temperature less than that of said particles.

16. A multi-zone element as defined in claim 12 wherein the particles of said particulate structure are solid spherical beads having a substantially uniform size within the range of from about 2 to 100 microns.

17. A multi-zone element for the analysis or transport of a liquid, said element comprising two or more zones in fluid contact, at least two of said zones having a particulate structure comprising:
(i) a plurality of heat-stable, organo-polymeric particles non-swellable in and impermeable to said liquid, said particles having a particle size of from about 1 to 200 microns, and (ii) an adhesive, in an amount less than 10 percent by weight of said particles, comprising an organic polymer different from that of said particles and insoluble in said liquid;
said adhesive being concentrated at contiguous surface areas of adjacent particles and bonding said particles into a coherent, three-dimensional lattice which is non-swellable in said liquid and which has interconnected void spaces among said particles to provide transport of said liquid, said lattice having a void volume of from about 25 to 80 percent.

18. A multi-zone element for the analysis of a liquid, said element comprising, in fluid contact, a zone having a particulate structure and a reagent zone containing one or more components of an interactive composition for said analysis, said particulate structure comprising:
(i) a plurality of heat-stable, organo-polymeric particles non-swellable in and impermeable to said liquid, said particles having a particle size of from about 1 to 200 microns, and (ii) an adhesive, in an amount less than 10 percent by weight of said particles, comprising an organic polymer different from that of said particles and insoluble in said liquid;
said adhesive being concentrated at contiguous surface areas of adjacent particles and bonding said particles into a coherent, three-dimensional lattice which is non-swellable in said liquid and which has interconnected void spaces among said particles to provide transport of said liquid, said lattice having a void volume of from about 25 to 80 percent.

19. A particulate structure for the analysis or transport of a liquid comprising:
(a) a plurality of heat-stable, organo-polymeric particles non-swellable in and impermeable to water, said particles having a particle size of from about 1 to 200 microns, and (b) an adhesive, in an amount less than 10 percent by weight of said particles, comprising an organic polymer different from that of said particles and insoluble in water;
said adhesive being concentrated at contiguous surface areas of adjacent particles and bonding said particles into a coherent, three-dimensional lattice which is non-swellable in water and which has interconnected void spaces among said particles to provide transport of said liquid, said lattice having a void volume of from about 25 to 80 percent.

20. A particulate structure for the analysis or transport of a liquid as defined in claim 19 wherein said organic polymer of said adhesive has a glass transition temperature less than that of said particles.

21. A particulate structure for the analysis of a liquid as defined in claim 19 wherein said particulate structure contains one or more components of an interactive composition for said analysis.

22. A particulate structure for the analysis of a liquid as defined in claim 19 wherein said particulate structure contains an immunoreagent.

23. A particulate structure as defined in claim 19 wherein said particles are solid particles of substantially uniform size.

24. A particulate structure as defined in claim 19 wherein the organic polymer of said particles and said adhesive comprise an addition copolymer of two or more different addition polymerizable monomers, at least one of said addition polymerizable monomers of said particles being common to at least one of said addition polymerizable monomers of said adhesive.

25. A particulate structure as defined in claim 19 wherein said particles contain a colorant.

26. An element for the analysis or transport of a liquid, said element comprising a radiation-transmissive support bearing a particulate structure comprising:
(i) a plurality of heat-stable, organo-polymeric particles non-swellable in and impermeable to water, said particles having a particle size of from about 1 to 200 microns, and (ii) an adhesive, in an amount less than 10 percent by weight of said particles, comprising an organic polymer different from that of said particles and insoluble in water;
said adhesive being concentrated at contiguous surface areas of adjacent particles and bonding said particles into a coherent, three-dimensional lattice which is non-swellable in water and which has interconnected void spaces among said particles to provide transport of said liquid, said lattice having a void volume of from about 25 to 80 percent.

27. A multi-zone element for the analysis or transport of a liquid, said element comprising, in fluid contact, a zone having a particulate structure and at least one other zone permeable to said liquid, said particulate structure comprising:
(i) a plurality of heat-stable, organo-polymeric particles non-swellable in and impermeable to water, said particles having a particle size of from about 1 to 200 microns, and (ii) an adhesive, in an amount less than 10 percent by weight of said particles, comprising an organic polymer different from that of said particles and insoluble in water;
said adhesive being concentrated at contiguous surface areas of adjacent particles and bonding said particles into a coherent, three-dimensional lattice which is non-swellable in water and which has interconnected void spaces among said particles to provide transport of said liquid, said lattice having a void volume of from about 25 to 80 percent.

28. A multi-zone element for the analysis of a liquid as defined in claim 27 wherein one or more components of an interactive composition for said analysis is located in at least one of said zones.

29. A multi-zone element for the analysis of a liquid as defined in claim 27 wherein at least one of said zones contains an immunoreagent.

30. A multi-zone element as defined in claim 27 wherein the particles of said particulate structure are solid spherical beads having a substantially uniform size within the range of from about 2 to 100 microns.

31. A multi-zone element for the analysis of a liquid, said element comprising, in fluid contact, a zone having a particulate structure and a reagent zone containing one or more components of an interactive composition for said analysis, said particulate structure comprising:
(i) a plurality of heat-stable, organo-polymeric particles non-swellable in and impermeable to water, said particles having a particle size of from about 1 to 200 microns, and (ii) an adhesive, in an amount less than 10 percent by weight of said particles, comprising an organic polymer different from that of said particles and insoluble in water;
said adhesive being concentrated at contiguous surface areas of adjacent particles and bonding said particles into a coherent, three-dimensional lattice which is non-swellable in water and which has interconnected void spaces among said particles to provide transport of said liquid, said lattice having a void volume of from about 25 to 80 percent; said particles comprising an addition polymer of an addition polymerizable blend of monomers, the total mono-mer content of said blend comprising at least one of the following monomers in the proportions indicated:
(a) from 0 to 100 weight percent of a polymerizable, amino-substituent-free styrene monomer;
(b) from 0 to about 25 weight percent of a polymer-izable acrylic ester;

(c) from 0 to 100 weight percent of a polymerizable methacrylic ester;
(d) from 0 to about 30 weight percent of a carboxylic acid containing one or more polymerizable ethylen-ically unsaturated groups;
(e) from 0 to about 75 weight percent of a nitrile containing one or more polymerizable ethylenically unsaturated groups, (f) from 0 to about 20 weight percent of a polymer-izable amine-substituted styrene monomer;
(g) from 0 to about 20 weight percent of an addition polymerizable monomer containing a crosslinkable group;
(h) from 0 to about 20 weight percent of a polymer-izable tertiary aminoalkyl acrylate or methacry-late;
(i) from 0 to 100 weight percent of a polymerizable, N-heterocyclic vinyl monomer;
(j) from 0 to about 20 weight percent of a polymer-izable acrylamide or methacrylamide monomer; and (k) from 0 to about 20 weight percent of a cross-linking monomer containing at least two addition polymerizable groups.

32. An element for the analysis or transport of an aqueous liquid, said element comprising a support bearing a particulate structure comprising:
(i) a plurality of heat-stable, organo-polymeric particles non-swellable in and impermeable to said liquid, said particles having a particle size of from about 1 to 200 microns, and (ii) an adhesive, in an amount of from about 1 to less than 10 percent by weight of said particles, comprising an organic polymer different from that of said particles and insoluble in said liquid;
said adhesive being concentrated at contiguous surface areas of adjacent particles and bonding said particles into a coherent, three-dimensional lattice which is non-swellable in said liquid and which has interconnected void spaces among said particles to provide transport of said liquid, said lattice having a void volume of from about 25 to 80 percent; said particles comprising an addition polymer of an addition polymerizable blend of monomers, the total monomer content of said blend comprising at least one of the follow-ing monomers in the proportions indicated:
(a) from 0 to 100 weight percent of a polymerizable, amino-substituent-free styrene monomer;
(b) from 0 to about 25 weight percent of a polymer izable acrylic ester;
(c) from 0 to 100 weight percent of a polymerizable methacrylic ester;
(d) from 0 to about 30 weight percent of a carboxylic acid containing one or more polymerizable ethyl-enically unsaturated groups;
(e) from 0 to about 75 weight percent of a nitrile containing one or more polymerizable ethylenically unsaturated groups;
(f) from 0 to about 20 weight percent of a polymer-izable amine-substituted styrene monomer;
(g) from 0 to about 20 weight percent of an addition polymerizable monomer containing a crosslinkable group;
(h) from 0 to about 20 weight percent of a polymer-izable tertiary aminoalkyl acrylate or methacry-late;
(i) from 0 to 100 weight percent of a polymerizable, N-heterocyclic vinyl monomer;
(j) from 0 to about 20 weight percent of a polymer-izable acrylamide or methacrylamide monomer; and (k) from 0 to about 20 weight percent of a cross-linking monomer containing at least two addition polymerizable groups.

33. An element as defined in claim 32 wherein the organo-polymer of the particles in said particulate struc-ture is selected from the group consisting of:
1. Polystyrene 2. Poly(styrene-co-methacrylic acid) 3. Poly(vinyltoluene-co-p-t-butylstyrene-co-methacrylic acid) 4. Poly(vinyltoluene-co-p-t-butylstyrene-co-methacrylic acid-co-divinylbenzene) 5. Poly(methyl methacrylate) 6. Poly(styrene-co-vinylbenzyl chloride-methacrylic acid) 7. Poly(styrene-co-N,N,N-trimethyl-N-vinylbenzyl-ammonium chloride-co-methacrylic acid) 8. Poly(styrene-co-divinylbenzene) 9. Poly(styrene-co-butyl acrylate-co-methacrylic acid) 10. Poly(styrene-co-methacrylic acid-co-divinylbenzene) 11. Poly(vinylbenzyl chloride-co-methacrylic acid-co-divinylbenzene) 12. Poly(styrene-co-2-hydroxyethyl methacrylate-co-methacrylic acid) 13. Poly(methyl methacrylate-co-butyl acrylate) 14. Poly(styrene-co-acrylonitrile) 15. Poly(methyl methacrylate-co-N-(m- and p-vinylbenzyl)-N,N-dimethylamine hydrochloride-co-ethylene dimethacrylate) 16. Poly(methyl methacrylate-co-2-(N,N-diethylamino-ethyl methacrylate hydrochloride-co-ethylene dimethacrylate).

34. An element for the analysis or transport of an aqueous liquid, said element comprising a support bearing a particulate structure comprising:
(i) a plurality of heat-stable, organo-polymeric particles non-swellable in and impermeable to said liquid, said particles having a particle size of from 2 to 200 microns, and (ii) an adhesive, in an amount of from about 1 to less than 5 percent by weight of said particles, comprising an organic polymer different from that of said particles and insoluble in said liquid;
said adhesive being concentrated at contiguous surface areas of adjacent particles and bonding said particles into a coherent, three-dimensional lattice which is non-swellable in said liquid and which has interconnected void spaces among said particles to provide transport of said liquid, said lattice having a void volume of from about 25 to 80 percent; said particles comprising an addition copolymer of an addition polymerizable blend of two or more different monomers, the total monomer content of said blend comprising at least one of the following monomers in the proportions indicated:
(a) from 0 to 99 weight percent of a polymerizable, amino-substituent-free styrene monomer;
(b) from 0 to about 25 weight percent of a polymer-izable acrylic ester;
(c) from 0 to 75 weight percent of a polymerizable methacrylic ester;
(d) from 0 to about 30 weight percent of a carboxylic acid containing one or more polymerizable ethyl-enically unsaturated groups;
(e) from 0 to about 75 weight percent of a nitrile containing one or more polymerizable ethylenically unsaturated groups;
(f) from 0 to about 20 weight percent of a polymer-izable amine-substituted styrene monomer;
(g1) from 0 to about 10 weight percent of an addition polymerizable monomer containing a group cross-linkable by a gelatin hardener;
(g2) from 0 to about 10 weight percent of an addition polymerizable monomer containing a group cross-linkable by a diamine;
(h) from 0 to about 20 weight percent of a polymer-izable tertiary aminoalkyl acrylate or methacry-late;
(i) from 0 to 75 weight percent of a polymerizable, N-heterocyclic vinyl monomer;
(j) from 0 to about 20 weight percent of a polymer-izable acrylamide or methacrylamide monomer; and (k) from 0 to about 5 weight percent of a crosslinking monomer containing at least two addition polymer-izable groups.

35. An element as defined in claim 34, the adhesive contained in said particulate structure of said element comprising an addition copolymer of two or more different addition polymerizable monomers, at least one of the addition polymerizable monomers of the adhesive being common to one of the monomers of the addition copolymer of said particles.

36. An element as defined in claim 34, the adhesive in said particulate structure of said element being present in an amount of from about 1 to 4 percent by weight based on the weight of said particles.

37. An element as defined in claim 34 wherein the adhesive in said particulate structure has a glass transition temperature less than that of said particles.

38. An element for the analysis of an aqueous liquid as defined in claim 34 wherein said element contains one or more components Or an interactive composition for said analysis.

39. An element for the analysis of an aqueous liquid as defined in claim 34 wherein said element contains an immunoreagent.

40. An element as defined in claim 34 wherein the particles in said particulate structure are solid spherical beads having a substantially uniform size within the range of from about 2 to 100 microns.

41. An element as defined in claim 34, the adhesive in said particulate structure of said element comprising an addition copolymer of an addition polymerizable blend of monomers selected from the following group:
A. a monomer blend containing from about 1 to 35 weight percent of one or more of said group (a) monomers with the remainder of the blend com-prising addition polymerizable alkyl acrylates or methacrylates;
B. a monomer blend containing from about 20 to 95 weight percent of monomers selected from group (a), (b), (c), (g1), (g2), and (k) with the remainder of the monomer blend comprising one or more addition polymerizable monomers having an active hydrogen or salts thereof; and C. a monomer blend comprising from about 15 to 100 weight percent of one or more monomers selected from the group consisting of 1-vinylimidazole, vinylbenzyl alcohol, ethyl acrylate, or a monomer from group (j) with the remaining monomers of the blend comprising monomers from group (g1).

42. An element as defined in claim 34, the adhesive in said particulate structure of said element comprising an addition copolymer of an addition polymerizable blend of monomers selected from the following group:
A. a monomer blend containing from about 10 to 30 weight percent of one or more of said group (a) monomers with the remainder of the blend com-prising addition polymerizable alkyl acrylates or methacrylates; and B. a monomer blend containing from about 50 to 95 weight percent Or monomers selected from group (a), (b), (c), and (k) with the remainder of the monomer blend comprising one or more addition polymerizable monomers having an active hydrogen or salts thereof.

43. An element as defined in claim 34, the adhesive in said particulate structure of said element comprising an addition copolymer of an addition polymer-izable blend of monomers containing from about 50 to 95 weight percent of monomers selected from groups (a), (b), and (c) with the remainder of said blend comprising one or more addition polymerizable monomers having an active hydrogen or salt thereof selected from the group consisting of acrylic acid, methacrylic acid, sulfo- or sulfate-substituted monomers, and the alkali metal and ammonium salts of these monomers.

44. An element as defined in claim 34 wherein the adhesive in said particulate structure is selected from the group consisting of:
1. Poly(n-butyl acrylate-co-styrene-co-2-acrylamido-2-methylpropanesulfonic acid) 2. Poly(butyl acrylate-co-styrene-co-2-acrylamido-2-methylpropanesulfonic acid) 3. Poly(ethyl acrylate-co-acrylic acid-co-2-aceto-acetoxyethyl methacrylate-co-2-acrylamido-2-methyl-propanesulfonic acid) 4. Poly(vinylbenzyl alcohol) 5. Poly(ethyl acrylate) 6. Poly(N-isopropylacrylamide) 7. Poly(2-hydroxyethyl methacrylate-co-2-acetoacetoxy-ethyl methacrylate) 8. Poly(n-butyl acrylate-co-acrylic acid) 9. Poly(n-butyl acrylate-co-acrylic acid-co-methacrylic acid-co-ethyl acryloylacetate) 10. Poly(n-butyl acrylate-co-acrylic acid-co-ethyl acryloylacetate) 11. Poly(n-butyl acrylate-co-methacrylic acid-co-2-acetoacetoxyethyl methacrylate) 12. Poly(n-butyl acrylate-co-styrene) 13. Poly(n-butyl acrylate-co-2-acrylamido-2-methylpropane-sulfonic acid-co-2-acetoacetoxyethyl methacrylate) 14. Poly(n-butyl acrylate-co-acrylic acid-co-2-acetoacetoxy-ethyl methacrylate-co-2-acrylamido-2-methylpropane sulfonic acid) 15. Poly(n-butyl methacrylate-co-styrene) 16. Poly(ethyl acrylate-co-styrene) 17. Poly(n-butyl acrylate-co-2-acrylamido-2-methyl-propanesulfonic acid) 18. Poly(n-butyl acrylate-co-styrene) 19. Poly(2-ethylhexyl acrylate-co-acrylic acid-co-2-acetoacetoxyethyl methacrylate-co-2-acrylamido-2-methylpropanesulfonic acid) 20. Poly(n-butyl acrylate-co-methacrylic acid) 21. Poly(ethyl acrylate-co-acrylic acid) 22. Poly(butyl acrylate-co-styrene-co-2-acetoacetoxyethyl methacrylate-co-2-acrylamido-2-methylpropanesulfonic acid) 23. Poly(butyl acrylate-co-styrene-co-2-acrylamido-2-methylpropanesulfonic acid-co-divinylbenzene) 24. Poly(acrylamide-co-2-acetoacetoxyethyl methacrylate)-

45. A multilayer element for the analysis or transport of an aqueous liquid, said element containing two or more superposed layers permeable to said liquid in fluid contact with one another, one of said layers having a particulate structure comprising:
(i) a plurality of heat-stable, organo-polymeric particles non-swellable in and impermeable to said liquid, said particles having a particle size of from about 1 to 200 microns, and (ii) an adhesive, in an amount less than 10 percent by weight of said particles, comprising an organic polymer different from that of said particles and insoluble in said liquid;
said adhesive being concentrated at contiguous surface areas of adjacent particles and bonding said particles into a coherent, three-dimensional lattice which is non-swellable in said liquid and which has interconnected void spaces among said particles to provide transport of said liquid, said lattice having a void volume of from about 25 to 80 percent.

46. A multilayer element as defined in claim 45 wherein the organic polymer of the adhesive in said par-ticulate structure has a glass transition temperature less than that of said particles.

47. A multilayer element as defined in claim 45 wherein the particles of said particulate structure are solid spherical beads having a substantially uniform size within the range of from about 2 to 100 microns.

48. A multilayer element as defined in claim 45 wherein the organo-polymeric composition of the particles in said particulate structure comprises an organic polymer containing a repeating unit having a chemical group re-presenting an active bonding site for chemical attachment to an interactive composition for said analysis.

49. A multilayer element for the analysis of an aqueous liquid as defined in claim 45 wherein at least one of said layers contains an immunoreagent.

50. A multilayer element for the analysis of an aqueous liquid as defined in claim 45 wherein one or more components of an interactive composition for said analysis is located in at least one of said layers.

51. A multilayer element for the analysis or transport of an aqueous liquid, said element containing two or more superposed layers in fluid contact, at least two of said layers having a particulate structure comprising:
(i) a plurality of heat-stable, organo-polymeric particles non-swellable in and impermeable to said liquid, said particles having a particle size of from about 1 to 200 microns, and (ii) an adhesive, in an amount of from about 1 to less than 10 percent by weight of said particles, comprising an organic polymer different from that of said particles and insoluble in said liquid;
said adhesive being concentrated at contiguous surface areas of adjacent particles and bonding said particles into a coherent, three-dimensional lattice which is non-swellable in said liquid and which has interconnected void spaces among said particles to provide transport of said liquid, said lattice having a void volume of from about 25 to 80 percent; said particles comprising an addition polymer of an addition polymerizable blend of monomers, the total monomer content of said blend comprising at least one of the follow-ing monomers in the proportions indicated:
(a) from 0 to 100 weight percent of a polymerizable, amino-substituent-free styrene monomer;
(b) from 0 to about 25 weight percent of a polymer-izable acrylic ester;
(c) from 0 to 100 weight percent of a polymerizable methacrylic ester;
(d) from 0 to about 30 weight percent of a carboxylic acid containing one or more polymerizable ethyl-enically unsaturated groups;
(e) from 0 to about 75 weight percent of a nitrile containing one or more polymerizable ethylenically unsaturated groups;
(f) from 0 to about 20 weight percent of a polymer-izable amine-substituted styrene monomer;
(g) from 0 to about 20 weight percent of an addition polymerizable monomer containing a crosslinkable group;
(h) from 0 to about 20 weight percent of a polymer-izable tertiary aminoalkyl acrylate or methacry-late;
(i) from 0 to 100 weight percent of a polymerizable, N-heterocyclic vinyl monomer;
(j) from 0 to about 20 weight percent of a polymer-izable acrylamide or methacrylamide monomer; and (k) from 0 to about 20 weight percent of a cross-linking monomer containing at least two addition polymerizable groups.

52. A multilayer element as defined in claim 51 wherein the particles of said particulate structure comprise spherical beads having a uniform particle size within the range of from about 1 to 20 microns and wherein one of said layers having said particulate structure contains an immuno-reagent immobilized on said beads.

53. A multilayer element as defined in claim 51 wherein the particles of said particulate structure comprise spherical beads having a uniform particle size within the range of from about 1 to 20 microns and wherein one of said layers having said particulate structure contains an antibody immobilized on said beads.

54. A multilayer element as defined in claim 51 wherein the particles of said particulate structure comprise spherical beads having a uniform particle size within the range of from about 5 to 20 microns and wherein one of said layers having said particulate structure contains an anti-body immobilized on said beads.

55. A multilayer element for the analysis of an aqueous liquid, said element having a radiation-transmissive support bearing at least one superposed reagent layer permeable to said liquid, said reagent layer containing one or more components of an interactive composition for said analysis, and in fluid contact with and superposed over said reagent layer at least one layer having a particulate structure comprising:
(i) a plurality of heat-stable, organo-polymeric particles non-swellable in and impermeable to said liquid, said particles having a particle size of from about 1 to 200 microns, and (ii) an adhesive, in an amount of from about 1 to less than 10 percent by weight of said particles, comprising an organic polymer different from that of said particles and insoluble in said liquid;
said adhesive being concentrated at contiguous surface areas of adjacent particles and bonding said particles into a coherent, three-dimensional lattice which is non-swellable in said liquid and which has interconnected void spaces among said particles to provide transport of said liquid, said lattice having a void volume of from about 25 to 80 percent; said particles comprising an addition polymer of an addition polymerizable blend of monomers, the total monomer content of said blend comprising at least one of the follow-ing monomers in the proportions indicated:
(a) from 0 to 100 weight percent of a polymerizable, amino-substituent-free styrene monomer;
(b) from 0 to about 25 weight percent of a polymer-izable acrylic ester;
(c) from 0 to 100 weight percent of a polymerizable methacrylic ester;
(d) from 0 to about 30 weight percent of a carboxylic acid containing one or more polymerizable ethyl enically unsaturated groups;
(e) from 0 to about 75 weight percent of a nitrile containing one or more polymerizable ethylenically unsaturated groups;
(f) from 0 to about 20 weight percent of a polymer-izable amine-substituted styrene monomer;
(g) from 0 to about 20 weight percent of an addition polymerizable monomer containing a crosslinkable group;
(h) from 0 to about 20 weight percent of a polymer-izable tertiary aminoalkyl acrylate or methacry-late;
(i) from 0 to 100 weight percent of a polymerizable, N-heterocyclic vinyl monomer, (j) from 0 to about 20 weight percent of a polymer-izable acrylamide or methacrylamide monomer; and (k) from 0 to about 20 weight percent of a cross-linking monomer containing at least two addition polymerizable groups.

56. A multilayer element as defined in claim 55 wherein the particles in said layer having said particulate structure comprise solid, spherical beads having a uniform particle size within the range of from about 20 to 100 microns.

57. A multilayer element as defined in claim 55 wherein said interactive composition forms or releases a radiometrically detectable species.

58. A multilayer element as defined in claim 55 wherein said interactive composition forms or releases a colorimetrically or fluorometrically detectable species and wherein said element contains a registration layer to receive said detectable species, said registration layer intervening said support and said reagent layer.

59. A multilayer element as defined in claim 55 wherein (i) said interactive composition forms or releases a colorimetrically or fluorometrically detectable species, (ii) said element contains a registration layer to receive said detectable species, said registration layer intervening said support and said reagent layer, and (iii) said element contains a radiation-blocking agent to screen said detectable species in said regis-tration layer from one or more other layers of said element.

60. A multilayer element for the analysis of an antigen contained in an aqueous liquid, said element having a radiation-transmissive support bearing two or more super-posed layers in fluid contact, at least two of said layers having a particulate structure, the layer with said particu-late structure furthest removed from said support containing immobilized immunoreagent, said particulate structure comprising:
(i) a plurality of heat-stable, organo-polymeric particles non-swellable in and impermeable to said liquid, said particles having a particle size of from about 1 to 200 microns, and (ii) an adhesive, in an amount of from about 1 to less than 10 percent by weight of said particles, comprising an organic polymer different from that of said particles and insoluble in said liquid;
said adhesive being concentrated at contiguous surface areas of adjacent particles and bonding said particles into a coherent, three-dimensional lattice which is non-swellable in said liquid and which has interconnected void spaces among said particles to provide transport of said liquid, said lattice having a void volume of from about 25 to 80 percent; said particles comprising an addition polymer of an addition polymerizable blend of monomers, the total monomer content of said blend comprising at least one of the follow-ing monomers in the proportions indicated:
(a) from 0 to 100 weight percent of a polymerizable, amino-substituent-free styrene monomer;
(b) from 0 to about 25 weight percent of a polymer-izable acrylic ester;
(c) from 0 to 100 weight percent of a polymerizable methacrylic ester;
(d) from 0 to about 30 weight percent of a carboxylic acid containing one or more polymerizable ethyl-enically unsaturated groups;
(e) from 0 to about 75 weight percent of a nitrile containing one or more polymerizable ethylenically unsaturated groups;
(f) from 0 to about 20 weight percent of a polymer-izable amine-substituted styrene monomer;
(g) from 0 to about 20 weight percent of an addition polymerizable monomer containing a crosslinkable group;
(h) from 0 to about 20 weight percent of a polymer-izable tertiary aminoalkyl acrylate or methacry-late;
(i) from 0 to 100 weight percent of a polymerizable, N-heterocyclic vinyl monomer;
(j) from 0 to about 20 weight percent of a polymer-izable acrylamide or methacrylamide monomer; and (k) from 0 to about 20 weight percent of a cross-linking monomer containing at least two addition polymerizable groups.

61. A multilayer element as defined in claim 60 wherein the particles of said particulate structure com-prise spherical beads having a uniform particle size within the range of from about 1 to 20 microns and wherein said immunoreagent is antibody immobilized on said beads.

62. A multilayer element as defined in claim 60 wherein said immunoreagent is antibody and said layer con-taining said antibody contains a radiation-blocking agent.

63. A multilayer element as defined in claim 60 wherein said immunoreagent is antibody and said layer con-taining said antibody contains nonspecific protein.

64. A multilayer element as defined in claim 60 wherein said immunoreagent is antibody and the particles of said layer containing antibody contain a radiation-blocking agent.

65. A multilayer element as defined in claim 60 wherein said immunoreagent is antibody and said element con-tains an antigen bearing a label comprising a radiometrically detectable species.

66. A multilayer element as defined in claim 60 wherein said immunoreagent is antibody and said element contains an antigen bearing a label comprising a fluoro-metrically detectable species.

67. A multilayer element for the analysis of an antigen contained in an aqueous liquid, said element having a radiation-transmissive support bearing two or more super-posed layers in fluid contact, at least two of said layers having a particulate structure, the layer with said parti-culate structure furthest removed from said support con-taining immobilized antibody, said particulate structure comprising:
(i) a plurality of heat-stable, organo-polymeric particles non-swellable in and impermeable to said liquid, said particles having a particle size of from about 2 to 20 microns, and (ii) an adhesive, in an amount of from about 1 to less than 5 percent by weight of said particles, comprising an organic polymer different from that of said particles and insoluble in said liquid;
said adhesive being concentrated at contiguous surface areas of adjacent particles and bonding said particles into a coherent, three-dimensional lattice which is non-swellable in said liquid and which has interconnected void spaces among said particles to provide transport Or said liquid, said lattice having a void volume Or from about 25 to 80 percent; said particles comprising an addition polymer of an addition polymerizable blend of monomers, the total monomer content said blend comprising at least one of the following monomers in the proportions indicated:
(a) from 0 to 99 weight percent or a polymerizable, amino-substituent free styrene monomer;
(b) from 0 to about 25 weight percent of a polymer-izable acrylic ester;
(c) from 0 to 75 weight percent of a polymerizable methacrylic ester;
(d) from 0 to about 30 weight percent of a carboxylic acid containing one or more polymerizable ethyl enically unsaturated groups;
(e) from 0 to about 75 weight percent of a nitrile containing one or more polymerizable ethylenically unsaturated groups;
(f) from 0 to about 20 weight percent of a polymer-izable amine-substituted styrene monomer;
(g1) from 0 to about 10 weight percent of an addition polymerizable monomer containing a group cross-linkable by a gelatin hardener;
(g2) from 0 to about 10 weight percent of an addition polymerizable monomer containing a group cross-linkable by a diamine;
(h) from 0 to about 20 weight percent of a polymer-izable tertiary aminoalkyl acrylate or methacry-late;
(i) from 0 to 75 weight percent of a polymerizable, N-heterocyclic vinyl monomer;
(j) from 0 to about 20 weight percent of a polymer-izable acrylamide or methacrylamide monomer; and (k) from 0 to about 5 weight percent of a crosslinking monomer containing at least two addition polymer-izable groups.

68. A multilayer element as defined in claim 67 the adhesive in said particulate structure comprising an addition copolymer of an addition polymerizable blend of monomers selected from the following group:
A. a monomer blend containing from about 1 to 35 weight percent of one or more of said group (a) monomers with the remainder of the blend com-prising addition polymerizable alkyl acrylates or methacrylates;
B. a monomer blend containing from about 20 to 95 weight percent of monomers selected from group (a), (b), (c), (g1), (g2), and (k) with the remainder of the monomer blend comprising one or more addition polymerizable monomers having an active hydrogen or salts thereof; and C. a monomer blend comprising from about 15 to 100 weight percent of one or more monomers selected from the group consisting of 1-vinylimidazole, vinylbenzyl alcohol, ethyl acrylate, or a monomer from group (j) with the remaining monomers of the blend comprising monomers from group (g1).

69. A multilayer element as defined in claim 67, the adhesive in said particulate structures comprising an addition copolymer of an addition polymerizable blend of monomers selected from the following group:
A. a monomer blend containing from about 10 to 30 weight percent of one or more of said group (a) monomers with the remainder of the blend com-prising addition polymerizable alkyl acrylates or methacrylates; and B. a monomer blend containing from about 50 to 95 weight percent of monomers selected from group (a), (b), (c), and (k) with the remainder of the monomer blend comprising one or more addition polymerizable monomers having an active hydrogen or salts thereof.

70. A multilayer element as defined in claim 67, the adhesive in said particulate structure comprising an addition copolymer of an addition polymerizable blend of monomers containing from about 50 to 95 weight percent of monomers selected from groups (a), (b), and (c) with the remainder of said blend comprising one or more addition polymerizable monomers having an active hydrogen or salt thereof selected from the group consisting of acrylic acid, methacrylic acid, sulfo- or sulfate-substituted monomers, and the alkali metal and ammonium salts of these monomers.

71. A multilayer element as defined in claim 67, the organo-polymer in the particles of said particulate structure being poly(styrene-co-methacrylic acid) or poly(styrene-co-vinylbenzyl chloride-methacrylic acid), and the organic polymer of the adhesive of said particulate structures being poly(n-butyl acrylate-co-styrene-co-2-acrylamido-2-methylpropanesulfonic acid).

72. A multilayer element as defined in claim 67 which contains an antigen bearing a label, said label com-prising a polymeric latex bead containing a fluorescent rare earth chelate.

73. A multilayer element for the analysis of an antigen contained in an aqueous liquid, said element having a radiation-transmissive support having a monolayer of antibody adsorbed thereto and superposed over said monolayer a particulate structure comprising:
(i) a plurality of heat-stable, organo-polymeric particles non-swellable in and impermeable to said liquid, said particles having a particle size of from about 1 to 200 microns, and (ii) an adhesive, in an amount of from about 1 to less than 10 percent by weight of said particles, comprising an organic polymer different from that of said particles and insoluble in said liquid;
said adhesive being concentrated at contiguous surface areas of adjacent particles and bonding said particles into a coherent, three-dimensional lattice which is non-swellable in said liquid and which has interconnected void spaces among said particles to provide transport of said liquid, said lattice having a void volume of from about 25 to 80 percent; said particles comprising an addition polymer of an addition polymerizable blend of monomers, the total monomer of said blend comprising at least one of the following monomers in the proportions indicated:
(a) from 0 to 100 weight percent of a polymerizable, amino-substituent-free styrene monomer;
(b) from 0 to about 25 weight percent of a polymer-izable acrylic ester;
(c) from 0 to 100 weight percent of a polymerizable methacrylic ester;
(d) from 0 to about 30 weight percent of a carboxylic acid containing one or more polymerizable ethyl-enically unsaturated groups;
(e) from 0 to about 75 weight percent of a nitrile containing one or more polymerizable ethylenically unsaturated groups;
(f) from 0 to about 20 weight percent of a polymer-izable amine substituted styrene monomer;
(g) from 0 to about 20 weight percent of an addition polymerizable monomer containing a crosslinkable group;

(h) from 0 to about 20 weight percent of a polymer-izable tertiary aminoalkyl acrylate or methacry-late;
(i) from 0 to 100 weight percent of a polymerizable, N-heterocyclic vinyl monomer;
(j) from 0 to about 20 weight percent of a polymer-izable acrylamide or methacrylamide monomer; and (k) from 0 to about 20 weight percent of a cross-linking monomer containing at least two addition polymerizable groups.

74. A multilayer element for the analysis of an antigen contained in an aqueous liquid, said element having a radiation-transmissive support bearing a scintillation layer and superposed thereover a particulate structure containing immobilized antibody, said particulate structure comprising:
(i) a plurality of heat-stable, organo-polymeric particles non-swellable in and impermeable to said liquid, said particles having a particle size of from about 1 to 200 microns, and (ii) an adhesive, in an amount of from about 1 to less than 10 percent by weight of said particles, comprising an organic polymer different from that of said particles and insoluble in said liquid;
said adhesive being concentrated at contiguous surface areas of adjacent particles and bonding said particles into a coherent, three-dimensional lattice which is non-swellable in said liquid and which has interconnected void spaces among said particles to provide transport of said liquid, said lattice having a void volume of from about 25 to 80 percent; said particles comprising an addition polymer of an addition polymerizable blend of monomers, the total monomer content of said blend comprising at least one of the follow-ing monomers in the proportions indicated:
(a) from 0 to 100 weight percent of a polymerizable, amino-substituent-free styrene monomer;
(b) from 0 to about 25 weight percent of a polymer-izable acrylic ester;
(c) from 0 to 100 weight percent of a polymerizable methacrylic ester;

(d) from 0 to about 30 weight percent of a carboxylic acid containing one or more polymerizable ethyl-enically unsaturated groups;
(e) from 0 to about 75 weight percent of a nitrile containing one or more polymerizable ethylenically unsaturated groups;
(f) from 0 to about 20 weight percent of a polymer-izable amine-substituted styrene monomer;
(g) from 0 to about 20 weight percent of an addition polymerizable monomer containing a crosslinkable group;
(h) from 0 to about 20 weight percent of a polymer-izable tertiary aminoalkyl acrylate or methacry-late;
(i) from 0 to 100 weight percent of a polymerizable, N-heterocyclic vinyl monomer;
(j) from 0 to about 20 weight percent of a polymer-izable acrylamide or methacrylamide monomer; and (k) from 0 to about 20 weight percent of a cross-linking monomer containing at least two addition polymerizable groups.

75. A multilayer element as defined in claim 74 wherein said scintillation layer comprises a particulate structure comprising:
(i) a plurality of heat-stable, organo-polymeric particles non-swellable in and impermeable to said liquid, said particles having a particle size of from about 1 to 200 microns, and (ii) an adhesive, in an amount less than 10 percent by weight of said particles, comprising a fluor-imbibed latex polymer different from that of said particles and insoluble in said liquid;
said adhesive being concentrated at contiguous surface areas of adjacent particles and bonding said particles into a coherent, three-dimensional lattice which is non swellable in said liquid and which has interconnected void spaces among said particles to provide transport of said liquid, said lattice having a void volume of from about 25 to 80 percent.

76. A multilayer element for the analysis of an antigen contained in an aqueous liquid, said element generating an enzyme-induced detectable change corresponding to the presence and/or concentration Or said antigen by interaction of said antigen, a separate quantity of the same antigen labelled with said enzyme, and antibody for said antigen;
said element having a radiation-transmissive support bearing two or more superposed layers in fluid contact including a reagent layer comprising at least a portion of an interactive enzyme assay composition which produces said enzyme-induced detectable change and superposed over said reagent layer a layer having a particulate structure containing immobilized antibody, said particulate structure comprising:
(i) a plurality of heat-stable, organo-polymeric particles non-swellable in and impermeable to said liquid, said particles having a particle size of from about 1 to 200 microns, and (ii) an adhesive, in an amount less than 10 percent by weight of said particles, comprising an organic polymer different from that of said particles and insoluble in said liquid;
said adhesive being concentrated at contiguous surface areas of adjacent particles and bonding said particles into a coherent, three-dimensional lattice which is non-swellable in said liquid and which has interconnected void spaces among said particles to provide transport of said liquid, said lattice having a void volume of from about 25 to 80 percent.

77. A method Or making an element for the analysis or transport of a liquid, said element containing a support bearing a particulate structure, said method comprising:
(a) forming in a liquid carrier a stable dispersion of (i) a plurality of heat-stable, organo-polymeric particles non-swellable in and impermeable to said liquid under analysis, said particles having a particle size of from about 1 to 200 microns, and (ii) an adhesive, in an amount less than 10 percent by weight of said particles, comprising an organic polymer, said adhesive polymer being different from that of said particles and being insoluble in said liquid under analysis;
and (b) applying said dispersion to said support and removing said liquid carrier at a temperature below the heat-stability temperature of said particles to concentrate said adhesive at con-tiguous surface areas of adjacent particles and bond said particles, in situ, on said support into a coherent, three-dimensional lattice which is non-swellable in said liquid under analysis, has interconnected void spaces among particles of said lattice to provide transport of said liquid under analysis, and has a void volume of from about 25 to 80 percent.

78. A method of making an element for the analysis or transport of an aqueous liquid, said element containing a support bearing a particulate structure, said method com-prising:
(a) forming in a liquid carrier a stable dispersion of (i) a plurality of heat-stable, organo-polymeric particles non-swellable in and impermeable to said liquid under analysis, said particles having a particle size of from about 1 to 200 microns, and (ii) an adhesive, in an amount less than 10 percent by weight of said particles, comprising an organic polymer, said adhesive polymer being different from that of said particles, being insoluble in said liquid under analysis, and having a glass transition temperature less than that of said particles; and (b) applying said dispersion to said support and removing said liquid carrier at a temperature above the glass transition temperature of said adhesive polymer but below the heat-stability temperature of said particles to concentrate said adhesive at contiguous surface areas of adjacent particles and bond said particles, in situ, on said support into a coherent, three-dimensional lattice which is non-swellable in said liquid under analysis, has interconnected void spaces among particles of said lattice to provide trans-port of said liquid under analysis, and has a void volume of from about 25 to 80 percent.

79. A method of making an element as defined in claim 78 wherein a surfactant is added to said liquid carrier during step (a) to facilitate formation of said stable dispersion.

80. A method of making an element as defined in claim 78 wherein formation Or said stable dispersion in step (a) is facilitated by matching the specific gravity of said heat-stable particles to that of said liquid carrier.

81. A method of making an element as defined in claim 78 wherein formation of said stable dispersion in step (a) is facilitated by matching the specific gravity of said heat-stable particles and that of said liquid carrier to a value within the range of from about 0.7 to 1.3.

82. A method of making an element as defined in claim 78 wherein formation of said stable dispersion in step (a) comprises:
preparing said adhesive as a latex, said latex comprising said adhesive polymer dispersed as a finely divided discontinuous phase in an aqueous liquid vehicle as a continuous phase; and admixing said heat-stable particles with said latex, said aqueous liquid vehicle of said latex serving as at least a portion of the liquid carrier for said stable dispersion.

83. A method for the analysis of an analyte contained in a liquid, said method comprising (a) contacting together said liquid and an element having a support bearing a particulate structure comprising (i) a plurality of heat-stable, organo-polymeric particles non-swellable in and impermeable to said liquid, said particles having a particle size of from about 1 to 200 microns 9 and (ii) an adhesive, in an amount less than 10 per-cent by weight of said particles, comprising an organic polymer different from that of said particles and insoluble in said liquid, said adhesive concentrated at contiguous surface areas of adjacent particles bonding said particles into a coherent, three-dimensional lattice non-swellable in said liquid with interconnected void spaces among particles of said lattice to provide trans-port of said liquid, said lattice having a void volume of from about 25 to 80 percent, the analyte, or a reaction product of said analyte, interact-ing with the element to produce a detectable change within the element; and (b) detecting said change to determine the presence and/or concentration of said analyte.

84. A method for the analysis of an analyte contained in a sample of whole blood, said method comprising, (a) contacting together said sample and an element having a support bearing a particulate structure comprising (i) a plurality of heat-stable, organo-polymeric particles non-swellable in and impermeable to said sample, said particles having a particle size of from about 20 to 100 microns, and (ii) an adhesive, in an amount less than 10 per-cent by weight of said particles, comprising an organic polymer different from that of said particles and insoluble in said sample, said adhesive concentrated at contiguous surface areas of adjacent particles bonding said particles into a coherent, three dimensional lattice non-swellable in said sample with interconnected void spaces among particles of said lattice to provide transport of said sample, said lattice having a void volume of from about 25 to 80 percent, the analyte, or a reaction product of said analyte, inter-acting with the element to produce a detectable change within the element; and (b) detecting said change to determine the presence and/or concentration of said analyte.

85. A method for the analysis of an analyte contained in a sample of whole blood as defined in claim 84 wherein said analyte is hemoglobin which physically interacts with said element to produce an absorption peak at 540 nm and wherein said 540 nm absorption peak is detected to determine the concentration of hemoglobin in said sample.

86. A method for the analysis of an analyte contained in a sample of whole blood as defined in claim 84 wherein said particulate structure includes an inter-active composition which interacts with said analyte, or said reaction product of the analyte, to produce a detectable change in said element.

87. A method for the analysis of an analyte contained in an aqueous liquid, said method comprising:
(a) contacting together said liquid and a multi-zone element having two or more zones in fluid contact, at least two of said zones having a particulate structure comprising (i) a plurality of heat-stable, organo-polymeric particles non-swellable in and impermeable to said liquid, said particles having a particle size of from about 1 to 200 microns, and (ii) an adhesive, in an amount less than 10 percent by weight of said particles, comprising an organic polymer different from that of said particles and insoluble in said liquid, said adhesive concentrated at contiguous surface areas of adjacent particles bonding said particles into a coherent, three-dimensional lattice non-swellable in said liquid with interconnected void spaces among particles of said lattice to provide transport of said liquid, said lattice having a void volume of from about 25 to 80 percent, said particles in the particulate structure of said element which first contacts said liquid having a larger particle size than the particles contained in said other particulate structure(s), at least one of said other particulate structures containing an inter-active composition for the analyte or a reaction product of said analyte;
a component of said liquid being retained in said particulate structure which first contacts said sample, and said analyte, or said reaction product of the analyte, interacting with said interactive composition contained in one of said other particulate structure(s) to produce a detectable change within said element; and (b) detecting said change to determine the presence and/or concentration of said analyte.

88. A method for the analysis of an analyte contained in a sample of whole blood, said Method comprising:
(a) contacting together said sample and a multilayer element having a radiation-transmissive support bearing two or more superposed layers in fluid contact, at least two of said layers having a particulate structure comprising (i) a plurality of heat-stable, organo-polymeric particles non-swellable in and impermeable to said sample, said particles having a particle size of from about 1 to 200 microns, and (ii) an adhesive, in an amount less than 10 percent by weight of said particles, comprising an organic polymer different from that of said particles and which is insoluble in said sample, said adhesive concentrated at contiguous surface areas of adjacent particles bonding said particles into a coherent, three-dimensional lattice which is non-swellable in said liquid with interconnected void spaces among particles of said lattice to provide transport of said liquid, said lattice having a void volume of from about 25 to 80 percent said particles in the particulate structure of said element which first contacts said sample having a particle size within the range of from about 20 to 200 microns and said particles in said other particulate structure(s) having a smaller particle size, at least one of said other particulate structure(s) containing an interactive composition for the analyte or a reaction product of said analyte;
the red blood cells of said sample being retained in said particulate structure which first contacts said sample, and said analyte, or said reaction product of said analyte, interacting with said interactive composition contained in one of said other particulate structure(s) to produce a detectable change within said element; and (b) detecting said change to determine the presence and/or concentration of said analyte.

89. A method for the analysis of an analyte contained in a liquid, wherein said analyte, or a reaction product of said analyte, interacts within an analytical element to produce a detectable change corresponding to the presence and/or concentration of said analyte, said method comprising (a) providing as said analytical element an element having a support bearing a particulate structure comprising (1) a plurality of heat-stable, organo-polymeric particles non-swellable in and impermeable to said liquid, said particles having a particle size of from about 1 to 200 microns, and (ii) an adhesive, in an amount less than 10 per-cent by weight of said particles, comprising an organic polymer different from that of said particles and insoluble in said liquid, said adhesive concentrated at contiguous surface areas of adjacent particles bonding said particles into a coherent, three-dimensional lattice non-swellable in said liquid with interconnected void spaces among particles of said lattice to provide transport of said liquid, said lattice having a void volume of from about 25 to 80 percent;
and;
(b) contacting together said element and said liquid to interact said analyte, or a reaction product of said analyte, with said element and produce said detectable change.

90. A method for the analysis of an analyte con-tained in a liquid, said method comprising (a) providing as said analytical element an element having a support bearing a particulate structure comprising (i) a plurality of heat-stable, organo-polymeric particles non-swellable in and impermeable to said liquid, said particles having a particle size of from about 1 to 200 microns, and (ii) an adhesive, in an amount less than 10 percent by weight of said particles, comprising an organic polymer different from that of said particles and insoluble in said liquid, said adhesive concentrated at contiguous surface areas of adjacent particles bonding said particles into a coherent, three dimensional lattice non-swellable in said liquid with interconnected void spaces among particles of said lattice to provide transport of said liquid, said lattice having a void volume of from about 25 to 80 percent;

(b) contacting together said element and said liquid to interact said analyte, or a reaction product of said analyte, with said element and produce a detectable change within said element; and (c) detecting said change to determine the presence and/or concentration of said analyte.

91. A method for the analysis of an antigen contained in an aqueous liquid wherein said antigen, the same antigen bearing a detectable label, and an antibody for said antigen interact within an immunoassay element to produce a detectable change corresponding to the presence and/or concentration of said antigen, said method comprising (a) providing as said immunoassay element a multi-zone element having two or more zones in fluid contact, at least one of said zones containing antibody immobilized therein and at least one of said zones having a particulate structure comprising:
(i) a plurality of heat-stable, organo-polymeric particles non-swellable in and impermeable to said liquid, said particles having a particle size of from about 1 to 200 microns, and (ii) an adhesive, in an amount less than 10 percent by weight of said particles, com-prising an organic polymer different from that of said particles and insoluble in said liquid, said adhesive being concentrated at contiguous surface areas of adjacent particles and bonding said particles into a coherent, three-dimensional lattice which is non-swellable in said liquid and which has interconnected void spaces among said particles to provide transport of said liquid, aid lattice having a void volume of from about 25 to 80 percent; and (b) contacting together said aqueous liquid and said multi-zone element in the presence of a known quantity of antigen bearing a detectable label to form in the zone of said element containing said antibody an antigen-antibody complex comprising said antigen contained in said aqueous liquid, a portion of said known quantity of antigen bearing said detectable label, and said antibody while the remaining portion of antigen bearing said detect-able label migrates to one of said other zones, the formation of said complex producing (i) a detectable change in said zone containing said complex corresponding to said portion of antigen bearing said detectable label contained in said complex, or (ii) a detectable change in one of said other zones corresponding to the difference between the portion of antigen bearing said detectable label contained in said complex and that contained in said known quantity of antigen bearing said detectable label, both of said detectable changes (b)(i) and (b)(ii) also corresponding to the presence and/or concentration of said antigen contained in said aqueous liquid.

92. A method for the analysis of an antigen contained in an aqueous liquid wherein said antigen, the same antigen bearing a detectable label, and an antibody for said antigen interact within an immunoassay element, said method comprising (a) providing as said immunoassay element a multi-zone element having two or more zones in fluid contact, at least one of said zones containing antibody immobilized therein and at least one of said zones having a particulate structure comprising:
(i) a plurality of heat-stable, organo-polymeric particles non-swellable in and impermeable to said liquid, said particles having a particle size of from about 1 to 200 microns, and (ii) an adhesive, in an amount less than 10 percent by weight of said particles, com-prising an organic polymer different from that of said particles and insoluble in said liquid, said adhesive being concentrated at contiguous surface areas of adjacent particles and bonding said particles into a coherent, three-dimensional lattice which is non-swellable in said liquid and which has interconnected void spaces among said particles to provide transport of said liquid, said lattice having a void volume of from about 25 to 80 percent, (b) contacting together said aqueous liquid and said multi-zone element in the presence of a known quantity of antigen bearing a detectable label to form in the zone of said element containing said antibody an antigen-antibody complex comprising said antigen contained in said aqueous liquid, a portion of said known quantity of antigen bearing said detectable label, and said antibody while the remaining portion of antigen bearing said detect-able label migrates to one of said other zones, the formation of said complex producing (i) a detectable change in said zone containing said complex corresponding to said portion of antigen bearing said detectable label contained in said complex, or (ii) a detectable change in one of said other zones corresponding to the difference between the portion of antigen bearing said detectable label contained in said complex and that contained in said known quantity of antigen bearing said detectable label; and (c) detecting either of said changes to determine the presence and/or concentration of antigen in said aqueous liquid.

93. A method for the analysis of an antigen contained in an aqueous liquid as defined in claim 92 wherein said detectable label comprises a polymeric latex bead containing a fluorescent rare earth chelate.