WO2000075632A1

WO2000075632A1 - Disposable sample holder

Info

Publication number: WO2000075632A1
Application number: PCT/GB2000/002154
Authority: WO
Inventors: Michael Human; Richard John Dickinson
Original assignee: Biochrom Limited
Priority date: 1999-06-03
Filing date: 2000-06-05
Publication date: 2000-12-14
Also published as: GB9912790D0

Abstract

Disclosed is disposable sample holder suitable for use in ultra violet (UV) spectrophotometry, the sample holder comprising a body portion which is substantially opaque to UV radiation, and a window portion which is substantially transmissive of UV radiation, the window portion being positioned so as to be aligned with UV radiation incident upon the sample holder when the sample holder is placed in a spectrophotometer.

Description

Title: Disposable Sample Holder

Field of the Invention

This invention relates to disposable sample holders for use in spectrophotometry, and to a method of making or using such sample holders.

Background of the Invention

Spectrophotometry is a widely-used and well-known technique for analysis in quality control/assurance and research: samples of interest are placed in a sample holder or "cuvette", and the cuvette is positioned in a spectrophotometer in which light is directed through the cuvette containing the sample. The amount of light absorbed by the sample can be determined to provide information concerning the sample, typically regarding the concentration of a substance therein.

For many samples, analysis can be performed using light of visible wavelengths (around 350- 800nm). However, for some samples it is necessary to conduct spectrophotometric analysis using ultraviolet light (of wavelengths around 200-350nm). This applies particularly to analysis of samples where it is desired to determine the presence and/or concentration of nucleic acids (such as DNA, RNA and "peptide nucleic acid" or PNA) of various sizes.

In order to minimise cost, spectrophotometry cuvettes are generally made of cheap materials, such as synthetic plastics (e.g. polystyrene or polymethacrylate). These are suitable for use in spectrophotometry at visible wavelengths, but are not suitable for UV spectrophotometry as they transmit very little, if any, light of less than 325 nm wavelength. Accordingly, for UV spectrophotometry it is conventional to use a cuvette composed of pure quartz glass. This material is UV transmissible, but is very expensive, and difficult to machine. As a result, such cuvettes are not essentially 'disposable' and must be cleaned and re-used.

Moreover, many fields of activity require spectrophotometric analysis using very small sample volumes (typically 6-250 μl). This applies particularly in respect of measurement of samples comprising nucleic acids. Sample holders for use with such small volume samples are referred to as microcuvettes. Microcuvettes for UV spectrophotometry are disproportionately expensive, requiring manufacture to fine tolerances and use of expensive materials. Moreover, as may be imagined, cleaning of microcuvettes (to avoid cross-contamination during subsequent re-use) is laborious and time-consuming.

The present invention aims to provide a cheap sample holder for use in UV spectrophotometry, which may be disposed of after use, thereby avoiding the need for tedious cleaning procedures .

Summary of the Invention

In a first aspect the invention provides a disposable sample holder suitable for use in ultra violet (UV) spectrophotometry, the sample holder comprising a body portion which is substantially opaque to UV radiation, and a window portion which is substantially transmissive of UV radiation, the window portion being positioned so as to be aligned with UV radiation incident upon the sample holder when the sample holder is placed in a spectrophotometer .

In a second aspect the invention provides a method of making a disposable sample holder for use in UV spectrophotometry, the method comprising the steps of forming a body portion of the sample holder, which body portion is substantially opaque to UV radiation; and providing the sample holder with a window portion which is substantially transmissive of UV radiation, the window portion being positioned so as to be aligned with UV radiation incident upon the sample holder when the sample holder is placed in a spectrophotometer.

The sample holder of the invention conveniently takes the form of a cuvette, especially a microcuvette, and is advantageously shaped and dimensioned to be accommodated in the sample carriage of a conventional spectrophotometer. The normal maximum dimensions of a standard cuvette are about 12.5mm wide, by 12.5mm deep, by 45mm tall, and a sample holder in accordance with the invention will advantageously possess similar or identical maximum dimensions. The sample holder will conveniently be adapted for use with small sample volumes (i.e. in the range 6-250μl, preferably 6-50μl ).

The body portion will advantageously be formed from a relatively cheap material, especially a moulded synthetic plastics material (for example the polystyrene or polymethacrylate materials used in disposable cuvettes for spectrophotometry at visible light wavelengths), or, alternatively, a material opaque to visible wavelengths, such as polycarbonate, polybutylene terephthalate (PBT), or acetal. Alternatively, the body portion may be formed from a material which normally is UV transmissive, but is formulated so as to comprise a substance which renders the material UV opaque (and typically opaque to visible wavelengths also). The window portion will generally have a front face and a rear face, each face having a inner and outer surface. In a preferred embodiment the sample holder comprises a sample receiving portion within which at least some of the sample is located when spectrophotometric readings are being obtained. Conveniently the opposed ends of the sample receiving portion are formed and defined by the front and rear faces of the window portion. The sample receiving portion will normally take the form of a narrow channel. The side walls of the channel may be formed by the window portion or by the body portion.

Conveniently the body and window portions will form integral parts of the sample holder.

The front and rear faces of the window portion are aligned, such that a beam of UV light incident normally upon the front face will generally pass through the sample receiving portion and exit the sample holder via the rear face of the window portion and then be detected by a light detecting device in the spectrophotometer. This passage of light through the sample holder (and any sample therein) is generally referred to as the "light path" . For a number of reasons, it is desirable that the distance of the light path through the sample (the "path length") is known accurately. Conventionally, cuvettes are designed such that the path length is 10mm. Accordingly, in a sample holder in accordance with the invention, the inner surfaces of the front and rear faces of the window portion will generally be separated by a sample receiving portion of 10mm, but those skilled in the art will appreciate that the sample holder may be constructed to have any desired path length (as long as the path length is known to a high degree of accuracy e.g. +/- 0.1mm or less).

For present purposes, the body portion of the sample holder is considered "substantially opaque" to UV radiation if it has no more than half of the UV transmissibility of the window portion. Typically the body portion will transmit less than 5 % of light of any wavelength in the range 200-350nm, and preferably also transmit less than 5% of wavelengths 350-1 lOOnm incident upon it (which ranges include the UV and visible parts of the spectrum respectively).

As explained above, the window portion is substantially transmissive of UV radiation. For present purposes, the window portion is considered "substantially transmissive" of UV radiation if each face of thereof transmits at least 10% , preferably at least 35%, and more preferably at least 45%, of light (with a wavelength of 260-280nm) incident upon it. Accordingly, in preferred embodiments, the window portion of the sample holder as a whole will transmit at least 30% of UV light incident upon it (disregarding the effects of any scattering or absorption caused by substances present in the sample receiving portion), such that enough UV light is transmitted to allow an absorbance measurement to be made.

The window portion may comprise quartz glass, of the type conventionally used to produce UV spectrophotometry cuvettes. In other embodiments, the window portion may more conveniently comprise one or more moulded synthetic plastics materials, such as, but not limited to: cyclo-olefin copolymers (e.g. preferably amorphous, metallocene thermoplastic copolymers such as "Topas"™- which is an especially preferred material, manufactured by Ticona and obtainable from CTP Plasro), or specially formulated preparations of polymethylmethacrylate (PMMA) such as Lucry.TM (obtainable from BASF) and Plexiglas^R™ (obtainable from Habs).

Alternatively or additionally, the window portion may comprise a thin film of material such as polymethylpentene (PMP or TPX [TPX is a trademark of Mitsui Petrochemicals Ltd]) obtainable from Mitsui, polythene (PE) obtainable from Moore & Buckle, or co-extruded polypropylene (e.g. PP S2G polypropylene manufactured by ICI and obtainable from Soretrac).

The window portion may take the form of a continuous piece of substantially UV-transmissive material running around the sample holder which, preferably, is masked by UV opaque material other than at the opposed faces defining the opposed ends of the sample receiving portion. Alternatively the front and rear faces of the window portion may comprise discontinuous, discrete pieces of substantially UV-transmissive material, such as thin slabs or disks accommodated within apertures, gaps or recesses provided in the body portion. The slabs or disks may be held in place by adhesives, lugs, seals or any other suitable arrangement.

In those embodiments where the window portion comprises a thin film material, the film may conveniently be additionally or alternatively held in place by clips which cut through the film and which may, for example, enter into frictional engagement with the body portion. Clips, lugs, seals and the like, as aforementioned, will typically be formed from UV opaque materials and may thus be used to define the limits of the substantially UV-transmissive window portion.

It will be apparent to the person skilled in the art that the window portion will desirably be substantially transmissive of light of visible and near infra red (e.g. 280-700nm) wavelengths, so as to be suitable for use in spectrophotometric analysis at these wavelengths, as well as at UV (260-280nm) wavelengths.

In a preferred embodiment at least one of the window portion and the body portion is formed from a resiliently deformable material. Conveniently both the window portion and the body portion are formed from substantially the same (preferably resiliently deformable) material, with the exception that the material forming the body portion additionally comprises one or more compounds which render the body portion UV opaque, and preferably opaque to light of visible wavelengths also. Conveniently, the window portion is formed from clear Topas or other substantially UV-transmissive synthetic plastics material, and the body portion is formed from the same material but comprises a compatible UV- and visible light opacifier, such as a dyed polyolefin. Desirably, the window portion and body portion are formed (e.g. by moulding) as two separate components, and then joining the components, typically with a snap-fit engagement action although other types of engagement (e.g. screw-threaded) may be envisaged. Accordingly it is preferred that one portion is provided with a resiliently deformable latch member which may be received and retained within a co-operating detent provided in the other portion when the two portions are engaged. If desired, each portion may be provided with such a latch member, and such a detent to receive and retain the latch member of the other portion.

Generally, the window portion will be shaped and dimensioned such that at least part of the window portion will be received within the body portion.

A further feature of a preferred embodiment calls for two apertures to be provided in opposed faces of the body portion, the apertures being aligned at opposite ends of the sample receiving portion of the sample holder, the arrangement being such that light (especially UV light) can pass through a first aperture in the UV-opaque body portion, pass through one end of the sample receiving portion (defined by a front face of the UV-transmissive window portion), and along the sample receiving portion and exit via the correspondingly aligned rear face of the window portion and second aperture in the body portion.

In those embodiments where the sample receiving portion is formed in part by two or more different components, the separate components will desirably form a water-tight seal. More preferably, the entire sample receiving portion will be formed from a single component, which avoids the need to form a water-tight seal.

Preferably the front and rear faces of the window portion define opposed ends of the sample receiving portion of the sample holder, and it is preferred that both front and rear faces (and additionally the sides also, in preferred embodiments) are formed from a single, continuous component.

A general advantage of sample holders in accordance with the invention is that they are immediately compatible for use with conventional spectrophotometers for use in UV spectrophotometry, and do not require the use of further extraneous components such as UV- masking adapters or the like.

In yet a further embodiment, the sample holder may take the form of a substantially UV- opaque "liner" positioned within a UV-transmissive receptacle, the "liner" portion being provided with aligned apertures, as described above, to allow UV light to pass through the sample receiving portion of the sample holder. The liner portion may conveniently be formed from a cheap, mouldable material, especially a synthetic plastics material such as those used to make disposable cuvettes for spectrophotometry at visible light wavelengths (e.g. polystyrene or polymethacrylate) or, alternatively materials opaque to visible light (e.g. polycarbonate, PBT or acetal). These materials are particularly suitable for producing the intricate components necessary for microcuvettes. The liner portion, in effect, defines the sides of the sample receiving portion of the sample holder, whilst the substantially UV transmissive parts of the receptacle portion define the ends of the sample receiving portion. In this way, the UV absorbance of the liner portion does not interfere with the ability to use the sample holder in UV spectrophotometry.

If desired (especially if the two portions of the sample holder are intended to be readily separable), sealing means may be provided to assist in formation of a liquid-tight seal between the adjacent surfaces of the respective portions. Thus, for example, parts of one and/or both portions may be provided with resiliently deformable sealing means, such as silicone rubber gaskets and the like, to prevent loss of sample when the sample is placed in the sample holder. (In general however, it will normally be preferred that the two portions of the sample holder will typically not be readily separable.)

Where a UV-opaque liner portion is employed it will conveniently be formed from a cheap material and may thus be discarded and a fresh liner portion inserted into the receptacle portion. Any of the previous sample remaining in the receptacle portion is unlikely to be of sufficient size as to affect subsequent absorbance measurements and, in any event, once the liner portion has been discarded any sample remaining in the receptacle portion can readily be removed, if desired, by a simple washing step which is accomplished far more readily than for a conventional microcuvette because, once the liner portion is removed, the receptacle portion has a larger volume and a simply-shaped inner surface. Thus depending on the embodiment, either the liner portion alone, or the sample holder in its entirety, may be substantially disposable.

The invention will now be further described by way of illustrative example and with reference to the accompanying drawings, in which:

Figure 1 is a perspective view of a sample holder in accordance with of the invention in the form of a microcuvette;

Figure 2 is a plan view, to a different scale, of the sample holder illustrated in Figure 1;

Figures 3 and 4 are a front view and side sectional view respectively of the sample holder shown in Figure 2;

Figures 5 and 6 are sectional views of part of alternative embodiments of a sample holder in accordance with the invention;

Figure 7 shows an exploded view of an alternative embodiment of a sample holder in accordance with the invention;

Figure 8 is an end elevation of a preferred embodiment of a sample holder in accordance with the invention;

Figure 9 is a longitudinal sectional view of the same embodiment along the line A-A' in Figure 8; and

Figure 10 is a longitudinal sectional view of the same embodiment, perpendicular to the line A-A'.

Referring to Figure 1 , a sample holder (denoted generally by reference numeral 10) in accordance with the invention is shaped and dimensioned so as to fit within the sample carriage of a conventional spectrophotometer. The maximum dimensions of the sample holder are about 12.5mm x 12.5mm x 45mm. The sample holder comprises a body portion 12, and a window portion comprising a front face 14a and an opposed face 14b (not visible in Figure 1).

The body portion 12 comprises a moulded synthetic plastics material which is opaque to UV radiation. The body portion comprises a substantially square-section upper part 12a and a lower part 12b of the body portion which has an H-section, which facilitates manufacture.

The front face 14a of the window portion comprises a thin (approx. 0.5mm), near-circular piece of pure quartz glass or other substantially UV-transmissive material, accommodated within a suitably shaped and dimensioned recess formed in the body portion 12. The front face 14a is about 6mm in diameter. The rear face 14b of the window portion is essentially identical to the front face 14a.

The body portion 12 of the sample holder is internally inwardly tapered so as to form, between the front and rear faces 14a, b of the window portion, a narrow channel of substantially uniform cross-section, which constitutes the sample receiving portion 16 of the sample holder. One end of the sample receiving portion 16 is visible in Figure 1.

The sample receiving portion 16 is more clearly illustrated in Figures 2-4. As is apparent from Figures 2 and 4, the sample receiving portion 16 comprises a widened throat section 18, centrally positioned. The throat section 18 can accommodate the tip of a micropipette or similar device for delivering a small volume (e.g. 6-50 μl) of a liquid sample into the sample receiving portion 16 of the sample holder. The thickness of the front and rear faces 14 a, b of the window portion is such that the sample receiving portion 16 provides a path length of 10mm ( + /- 0.05mm).

Figures 5 and 6 are sectional views illustrating alternative embodiments in assembled form (Figure 5) and prior to assembly (Figure 6). In the alternative embodiment illustrated in Figure 5 the front and rear faces 14a, b of the window portion comprise a thin film of substantially UV-transmissive synthetic plastics material (such as polythene or polymethylpentene). Each piece of thin film is held in place by a circular securing clip 20 which is accommodated within a reciprocal recess provided in the body portion. The securing clips 20 are formed from a material opaque to UV light. Accordingly, a centrally-positioned aperture 22 is provided in each clip 20, to allow UV light to pass through the front face 14a of the window portion, any sample present in the sample receiving portion 16, and exit the sample holder via the rear face 14b of the window portion.

The embodiment illustrated in Figure 6 (shown prior to assembly) is substantially identical to the embodiment shown in Figure 5, with the exception that the clips 20 are considerably thicker and, upon assembly, are intended partially or wholly to penetrate the thin film. The clip may be provided with one or more pointed projections, if desired, in order to assist penetration and gripping of the thin film.

Referring to Figure 7, a sample holder in accordance with the invention comprises a receptacle portion 30 and a liner portion 32, which are shown separated from each other in the Figure.

The outer receptacle portion 30 is shaped and dimensioned so as to be accommodated in the sample carriage of a conventional spectrophotometer. The receptacle portion 30 takes the general form of a square-section tube with a flat bottom, and is of substantially uniform UV- transmissible material, such as quartz glass or UV-transmissible pyrex.

The liner portion 32 is shaped and dimensioned so as to be received within the receptacle portion 30 and to form a flush fit therewith, and generally corresponds in shape to the receptacle portion 30. The liner portion 32 is formed from a synthetic moulded plastics material which is substantially opaque to UV and to visible light, such as polycarbonate, PBT or acetal. The floor of the liner portion 32 is raised above the bottom of the receptacle portion 30 on two "legs" 34. Formed in the floor of the liner portion 32 is an open-ended channel 36. When the liner portion 32 is inserted into the receptacle portion 30, open-ended channel 36 is converted into a closed-ended channel, the opposed ends of which are defined by the opposed first and second faces 30a, 30b of the receptacle portion 30. The closed-ended channel constitutes the sample receiving portion of the sample holder. The sample receiving portion is held at the appropriate height (so as to be illuminated by the UV light of the UV spectrophotometer) by the legs 34 of the liner portion 32. Thus, UV light normally incident upon face 30a of the receptacle portion 30, can pass along the sample receiving portion (through the UV transmissible material of the receptacle portion 30a, through the sample in the channel 36 in the liner portion 32) and exit the sample holder via opposed face 30b.

A preferred embodiment of the invention is illustrated in Figures 8-10.

Referring to Figure 8, which shows an end elevation, a sample holder in accordance with the invention comprises an upper, window portion 40, and a lower, body portion 42, of substantially similar external proportions. The overall dimensions of the sample holder are such as to be accommodated within the sample carriage of a conventional spectrophotometer. The window portion 40 is moulded from clear Topas™, a UV-transmissive synthetic plastics material. The body portion 42 is moulded from coloured Topas™ prepared by addition of a polyolefin containing a grey dye which is opaque both to UV and light of visible wavelength.

Front and rear faces of the body portion are each formed with a single rectangular aperture, the apertures in the front and rear faces being aligned. One of these apertures, 44, is apparent in Figure 8. The body portion 42 is open at its upper end, forming an open-topped box-like chamber, into which is inserted the lower end of the window portion 40. The lower end of the body portion 42 has a generally smooth rectangular outer profile, as does the sample holder as a whole. Internally, the body portion 42 is provided with a resiliently deformable latching member 46, apparent in Figures 9 and 10. A wedge shaped projection at the upper end of the latching member 46 engages with a co-operating detent 48 provided on a projection on the underside of the window portion 40. Accordingly, when the window portion 40 and body portion 42 are pushed together, during assembly of the sample holder, the latching member 46 is secured, in an over centre closure, by the detent 48 with a snap-fit action. This forces window portion 40 and body portion 42 tightly together, and assists in aligning precisely a sample receiving portion 50 of the window portion 40 with the two apertures 44 provided in the body portion 42.

The outer surface of the window portion 40 is smooth, so as to form a substantially continuous flush surface with the body portion 42 on the exterior, but the lower end is of slightly reduced dimension, so as to be received within the box-like chamber at the upper end of the body portion 42. The inner surface of the window portion 40, for the most part, follows the contours of the outer surface but is inwardly tapered at the lower end towards a narrow channel, which constitutes the sample receiving portion 50. Accordingly, the entire sample receiving portion is formed within a single component. The tapering of the sides of the window portion help ensure that a liquid sample placed within the window portion is directed down towards the sample receiving portion 50.

Accordingly, in use, a small volume sample deposited in the window portion 40 fills the sample receiving portion 50. When placed in a conventional spectrophotometer carriage, UV light may pass through the front aperture 44 in the UV opaque body portion 42, and then sequentially through the front face of the UV transmissive window portion 40, the sample in the sample receiving portion 50, the rear face of the window portion 40, and the rear aperture 44 in the body portion 42, whence it may be detected by the UV detecting apparatus of the spectrophotometer .

It will be apparent to those skilled in the art that embodiments other than those specifically described or illustrated may be made, so as to obtain the benefit of the present invention, and such alternative embodiments are included within the scope of the present invention.

Claims

1. A disposable sample holder suitable for use in ultra violet (UV) spectrophotometry, the sample holder comprising a body portion which is substantially opaque to UV radiation, and a window portion which is substantially transmissive of UV radiation, the window portion being positioned so as to be aligned with UV radiation incident upon the sample holder when the sample holder is placed in a spectrophotometer.

2. A sample holder according to claim 1, in the form of a cuvette shaped and dimensioned to be accommodated in the sample carriage of a conventional spectrophotometer.

3. A sample holder according to claim 1 or 2, wherein the body portion and/or the window portion comprises a moulded synthetic plastics material.

4. A sample holder according to any one of claims 1, 2 or 3, wherein the window portion comprises one or more of the following: a cyclo-olefin copolymer; a specially formulated preparation of substantially UV-transmissive PMMA; quartz glass; or a thin film comprising one or more of polymethylpentene, polythene, or polypropylene.

5. A sample holder according to any one of the preceding claims wherein the window portion and the body portion are formed from substantially similar material, with the exception that the body portion additionally comprises one or more compounds which render the body portion UV-opaque.

6. A sample holder according to any one of the preceding claims wherein the body portion comprises two aligned apertures in opposed faces thereof, so as to permit the passage of UV light through the body portion.

7. A sample holder according to any one of the preceding claims, wherein the sample holder comprises a sample receiving portion within which the sample is located during spectrophotometric analysis of the sample, opposed ends of the sample receiving portion being defined by front and rear faces of the window portion.

8. A sample holder according to any one of the preceding claims wherein the window portion is formed from a single component.

9. A sample holder according to any one of the preceding claims, wherein the body portion and window portion are formed separately and may be engaged by a snap-fit action.

10. A sample holder according to any one of the preceding claims, wherein at least one of the window or body portion comprises a resiliently deformable latching member which, when the two portions are engaged, is received and retained within a co-operating detent provided on the other portion.

11. A sample holder according to any one of the preceding claims which can be used in combination with a conventional spectrophotometer without requiring the use of an adapter or other such apparatus.

12. A disposable microcuvette suitable for use in UV spectrophotometry, in accordance with any one of the preceding claims.

13. A method of making a disposable holder for use in UV spectrophotometry, the method comprising the steps of forming a body portion of the sample holder, which body portion is substantially opaque to UV radiation; and providing the sample holder with a window portion which is substantially transmissive of UV radiation, the window portion being positioned so as to be aligned with UV radiation incident upon the sample holder when the sample holder is placed in a spectrophotometer.

14. A method according to claim 13, performance of which results in a sample holder according to any one of claims 1-12.

15. A method according to claim 13 or 14, wherein the window portion and body portion are both separately formed from a synthetic plastics material, and the two components are then assembled.

16. A method according to claim 15, wherein the window portion and body portion are assembled with a snap-fit action.

17. A sample holder substantially as hereinbefore described and with reference to the accompanying drawings.

18. A method of making a sample holder substantially as hereinbefore described and with reference to the drawings.