WO2009021964A2

WO2009021964A2 - Optical biochip platform with plasmonic structure

Info

Publication number: WO2009021964A2
Application number: PCT/EP2008/060621
Authority: WO
Inventors: Brian Maccraith; Michal Trnavsky
Original assignee: Dublin City University
Priority date: 2007-08-13
Filing date: 2008-08-13
Publication date: 2009-02-19
Also published as: WO2009021964A3

Abstract

Luminescent sensors which are configured such that operably plasmonic effects contribute to the output of the sensors are described. In a preferred arrangement there is provided a sensor having an optical element which operably effects generation of a luminescence signal for subsequent detection, the optical element being configured such that operably plasmonic effects contribute to at least one of the generation or capture of the luminescence signal. The invention also provides an optical biochip platform with plasmonic structure for detecting fluorescence or other luminescent signals and optical sensor configurations or systems that utilize an optical chip.

Description

OPTICAL BIOCHIP PLATFORM WITH PLASMONIC STRUCTURE

Field of the Invention

This present disclosure relates to luminescent sensors and particularly to luminescent sensors which are configured such that operably plasmonic effects contribute to the output of the sensors. In a preferred arrangement there is provided an optical sensor which operably provides an output signal resultant from an excitation of a detection moiety to generate a first signal and an enhancement of that first signal by surface plasmon coupled excitation effects. The invention also provides an optical biochip platform with plasmonic structure for detecting fluorescence or other luminescent signals and optical sensor configurations or systems that utilize an optical chip.

Background

Luminescent sensors which are based on detection of an optical signal emitted upon excitation of a detection moiety are known. Based on the specifics of the detection moiety chosen and the nature of the excitation source used, these sensors may be used to detect the presence of one or more analytes.

In designing such sensors it is important to ensure that the detected signal is representative of the actual analyte that is being investigated as opposed to being for example contaminated by background signals or the like. It is also important in many applications that the sensors are capable of detecting small concentrations of the desired analyte and in these scenarios the ability to provide highly sensitive sensor configurations is desirable.

There is therefore a need for improved luminescent sensors.

Summary

These and other problems are addressed by a luminescent sensor provided in accordance with the present teaching. Such a sensor includes an optical element having a recognition area coated with a semi-transparent metallic film and a detection or capture moiety responsive to the presence or otherwise of predetermined analytes. By exposing the optical element to an environment, one or more of the predetermined analytes will bind to the detection moiety to form a complex that is optically linked or coupled to the optical element. The complex provided by the combination of the capture moiety with the desired analyte may include a luminophore. In another arrangement, a further treatment of the capture moiety/analyte complex may be required so as to effect a binding of a luminophore thereto. An example of a typical complex will include a sandwich assay wherein the analyte being detected is provided between two antibodies in a complex structure.

Irrespective of the specific type of complex provided, subsequent excitation of that complex will result in plasmonic effects contributing to the output of the optical element. These effects may be the excitation of the luminophore through for example surface plasmon resonance and the efficient collection of generated luminescence through surface plasmon coupled emission. In a first arrangement, the metallic film is provided on an upper surface of the optical element and the excitation is from below; this serves to effect generation of a luminescence output through surface plasmon resonance, the luminescence signal generated being coupled back into the optical element through surface plasmon coupled emission. In another arrangement the excitation is from above the detection moiety and the plasmon effects provide for enhanced capture of the generated luminescence; a generated signal being coupled into the optical element by surface plasmon coupled emission.

As was mentioned above, excitation may be effected through exposure of the optical element to an optical excitation source which may be provided either above or below the optical element. In another arrangement excitation may be provided by forming a complex having an electro-chemi-luminescent tag and effecting a subsequent electro-chemi-luminescent excitation of that optical element.

The optical element is desirably substantially paraboloid in form, having side walls extending upwardly from a planar substrate. Such a structure may be considered as being the resultant surface obtained by revolving a parabola around its axis. In a preferred arrangement, the paraboloid, or other three dimensional form, has a truncated upper surface such that the side walls are separated at the top of the optical element by a substantially planar surface. Desirably the recognition area is provided on this upper planar surface of the optical element, separated from the substrate by the side walls of the optical element. Where two or more optical elements are provided in an array, desirably the plurality of optical elements share a common substrate. The optical element is desirably a solid structure configured to operate by total internal reflection in order to convert the strong intensity maximum emitted by the excited analyte/detection moiety from large surface angles into collimated and conveniently detectablejays along the optical axis. In this way, the optical element will desirably preferentially direct light that enters into the optical element from the recognition area at an angle greater than or equal to the critical angle of the macroscopic elements forming the sensing environment, downwardly through the optical element for detection below the optical element. The macroscopic elements defining the sensing environment are typically the materials used for fabrication of the optical element- such as glass or a polymer- and the aqueous environment where the sensor is used. For a glass water interface this is about 61 degrees.

An optical element that may usefully be employed is desirably formed of a polymer having a refractive index greater than that of water. In this way the optical element may typically be fabricated using injection moulding techniques. Useful polymers have a refractive index of about 1.45-1.65, an example of which being ZEONEX™ which has a refractive index of about 1.52.

By configuring the optical element to totally internally reflect light it is possible, when illuminating the optical element from below, to effect generation of evanescent waves along the boundary surface between optical element and the medium where it is located. These evanescent waves contribute to the generation of surface plasmon resonance which can be used to effect excitation of the detection moiety.

Suitable detection moieties are any molecules that specifically bind to an analyte of interest and include, for example, antibodies and oligonucleotides. Where a plurality of optical elements are provided, individual ones of the plurality may comprise the same or different detection moieties depending upon the application.

The detection moiety is desirably optically coupled to the optical element. By "optically coupled," when referring to the relationship between a moiety and an optical element, includes but is not limited to luminescent molecules directly bound to or adsorbed onto the optical element; luminescent molecules indirectly attached to the optical element through one or more linker molecules; luminescent molecules entrapped within a film (e.g., a polymer or sol-gel matrix) that is coated onto the optical element; a non-luminescent molecule that is capable of binding to a luminescent molecule of interest. A particular preferred type of optical coupling that may be employed within the context of the present teaching is a covalent attachment to the substrate. It will be understood that the detection moiety itself exhibits a preference for binding to specific target analytes within the detection medium- typically an aqueous medium. On binding of the target analyte to the detection moiety, the resultant structure or complex may in itself be capable of exhibiting luminescence on excitation. In another arrangement a secondary label may be required, typically a secondary antibody, to create the necessary structure capable of luminescence on excitation. The complexes that are optically coupled to the optical element may be configured to have distinguishable spectral emission properties such that the output of a first optical element may be spectrally distinguished from the output of a second optical element. BRIEF DESCRIPTION OF THE DRAWINGS

The objects and features of the present disclosure, which are believed to be novel, are set forth with particularity in the appended claims. The present disclosure, both as to its organization and manner of operation, together with further objectives and advantages, may be best understood by reference to the following description, taken in connection with the accompanying drawings as set forth below:

Figure IA shows an example of the angular distribution of radiation data of a plasmon-coupled fluorescence, with the specifics of the data illustrated being representative of an air/glass interface and SiO₂ lOOnm spacer layer.

Figure IB is a schematic showing the propagation of light into an optical element that does not have such a plasmonic enhancement, as is provided by the prior art.

Figure 1C shows is a schematic showing how plasmonic enhancement preferentially directs light within a prescribed angular distribution into the optical element, in accordance with the teaching of the present invention.

Figure 2 is a schematic of an optical element provided in accordance with the present teaching.

Figure 3 shows a sensor array including a plurality of optical elements.

Figure 4 is a schematic showing light paths within a structure such as provided in Figure 3.

Figure 5 is schematic showing selective excitation of the optical elements from above in a manner that allows for individual optical elements to be illuminated differently to other optical elements.

Figure 6 is a schematic showing how a broad illumination may be used to concurrently illuminate a plurality of optical elements in the same array.

Figure 7 is a schematic showing how the excitation source may be angularly offset from the optical array. Figure 8 is an example of an experimental chip having individual optical elements arranged to preferentially target specific analytes.

Figure 9 shows results from the use of the chip of Figure 8.

DETAILED DESCRIPTION OF EXEMPLARY EMBODIMENTS

In accordance with a first embodiment of the present invention, an array of optical elements are provided each of which is configured to provide for enhancement of the optical output through a combination of plasmonic effects including surface plasmon resonance and surface plasmon coupled emission. Such plasmonic enhancement may be effected by incorporating onto individual ones of the optical elements, a semistransparent metallic film and a spacer layer so as to provide means for enhanced excitation via surface plasmon resonance (SPR) and/or collection of the resultant signal using surface -plasmon coupled emission (SPCE). The combination of these two effects results in efficient suppression of unbound analyte molecules and minimised detection volume. Such an optical element may also be geometrically configured to preferentially discriminate the angular distribution of light propagating into the optical element, such as is provided by configurations that are optimized to take advantage of supercritical angle fluorescence phenomena. In such an arrangement the incorporation of a plasmonic enhancement phenomenon into a highly efficient collector of a surface-generated fluorescence from a biochip can provide highly efficient optical elements which have application in detection of small amounts of analytes. Each of the techniques involved act independently, without causing any limitation to each other, and an optical element as provided in accordance with the teaching of the present invention may provide a highly sensitive device useful in particular in biomedical diagnostic application.

By having a plurality of optical elements on a shared substrate, an optical biochip platform is provided that may be used in a wide variety of commercial applications that require a compact, highly sensitive device. The optical biochip platform is suitable for fast, accurate readout from micro-arrays with high sensitivity. No bioassay device based on conjunction of plasmon-enhanced fluorescence and efficient fluorescence collection is currently available.

An optical element that may be usefully employed within the context of the present disclosure is one which preferentially allows for propagation of light derived from plasmonic effects into the optical element for subsequent detection. By employing plasmonic effects it is possible to deliver a highly collimated light cone into the optical element. As shown in Figure IA, fluorescence emission 100 (fluorescence being a specific type of luminescence) into a substrate as derived from excitation of a luminescent source 105 is highly directional with respect to the polar angle of the substrate surface and the angular distribution of radiation is relatively narrow . It will be appreciated that the specific figures or values shown are related to the nature of the substrate and the medium within which it is placed. In this exemplary arrangement the generated fluorescence which is derived from excitation of a source on the surface of the substrate 120 propagates at a highly contained angle of about 62 degrees.

Figure IB shows in schematic form a prior art arrangement which is not configured to include enhanced plasmonic effects. In such an arrangement excitation of a source 105 effects generation of a fluorescent signal 125 that will propagate into the substrate at angles about the critical angle θ_c of the system. Figure 1C is similar to Figure IA but shows the comparable arrangement of Figure IB modified to provide for plasmonic effects. It will be appreciated that the resultant propagation- which may be traced back to a surface plasmon coupled emission effect (SPCE), is at an angle, Bm_1n, greater than the critical angle, θ_cπticai. As such, if the optical geometry of the optical element within which the fluorescence is introduced is selected to preferentially allow for propagation of light introduced at angles greater than the critical angle, i.e is of the type that is optimized for detection of light resultant from the super critical angle fluorescence (SAF) phenomena that such a geometry will also be suitable for detection of SPCE effects. The range of angles of SPCE is a sub-manifold of the SAF effects.

The present disclosure is directed to a design to optimize the performance of a luminescent, typically fluorescence-based, biosensor that integrates a plasmonic structure into a low-cost, mass-producible optical biochip. In a first arrangement a luminescent system is provided which couples an optical biochip contains an array of paraboloid or other suitably dimensioned optical elements with an excitation source. In use an optical element is provided in an environment or medium where potential analytes are to be found. This is typically an aqueous environment and operably could be provided as part of a flow through system. Target analytes within the medium will preferentially bind with or couple to the detection moiety so as to form a sandwich arrangement. This sample may be capable of luminescence in its own right or may require a secondary label such as an antibody which will then adhere to the detection moiety/analyte combination. Collectively these structure can be categorized as complexes having as a component element lumiphores or a luminophore or in more specific examples fiuorophores; the category indicating the ability to exhibit luminescence or fluorescence on excitation. In either scenario once the optical element has been exposed for a sufficient time period to allow the formation of such structures, subsequent excitation will provide for generation of a luminescence output which be detected to provide evidence of the presence of the analyte within the test medium. If the excitation source is an optical excitation source and is arranged below the optical element it is possible, in accordance with the present teaching to provide for generation of evanescent waves at the boundary between the optical element and the acqueous environment to provide for surface plasmon resonance which can be used to induce excitation of the sample.

A plasmonic structure comprising a semi-transparent metallic film and a spacer layer formed on an optical element substrate provides a means for enhanced excitation via surface plasmon resonance (SPR) and enhanced collection of that luminescence through surface-plasmon coupled emission (SPCE). The combination of these two effects results in efficient suppression of unbound analyte molecules and minimised detection volume. It will be appreciated that the presence of the metallic film is required for the generation of surface plasmons and the dielectric spacer layer above ensures that the requisite separation of the luminophore from the optical element substrate.

An example of an optical element geometry that may be usefully employed within the teaching of the present invention is described in US Application No. 11/431,349, MacCraith et al., which is incorporated herein by reference. In this disclosure a plurality of solid parabolic optical elements are provided on a shared substrate so as to provide a chip. The geometry of the individual parabolic or indeed paraboloid elements on the chip are designed to facilitate both efficient excitation of surface-bound analyte molecules and collection of surface-generated fluorescence emitted above the critical angle of respective dielectric interface (supercritical angle fluorescence).

An example of an optical element which is based on such a geometry but modified to provide plasmonic effects is shown in Figure 2. In this arrangement, an optical element 200 in the form of a solid paraboloid structure having side walls 205 extending upwardly from a planar substrate 210 is provided. A planar top surface 215 which separates the two side walls is provided. This planar surface is desirably parallel with the surface of the substrate. This optical element is shown as being bottom illuminated in that excitation light 220 enters the optical element through the substrate. A recognition area 216 is provided on the top or upper surface of the optical element.

As was mentioned above, the optical element 200 is desirably a solid structure configured to operate by total internal reflection in order to efficiently collect the strong intensity maximum emitted by the excited complex of the analyte/detection moeity and received into the optical element. In this arrangement, the generated fluorescence signal is desirably detected by a detector provided below the substrate. 210. Similarly to that described in US 11/431,349, an annular aperture 240 may be provided below a bottom surface 211 of the substrate 210 to ensure that light entering the detecting surface at an angle less than the critical angle impinges on the aperture. It will be understood that as surface plasmon coupled emission preferentially directs light having an angle greater than or equal to the critical angle, that the use of such an aperture is not essential.

In accordance with the present disclosure, the recognition area 216 is modified to provide for enhanced plasmonic effects. In this case, the recognition area of the parabolic elements is coated with a thin semi transparent metal film 250 (~50nm, e.g. silver, gold, aluminum) and a thin dielectric spacer layer 251 (oxide, polymer film, polyelectrolyte multilayer etc.) with required thickness and dielectric constants. A detection moiety 253 is optically coupled to the optical element and is capable of binding with or tagging to a suitable analyte. Suitable detection moieties are any molecules that specifically bind to an analyte of interest and include, for example, antibodies and oligonucleotides. Where a plurality of optical elements are provided, individual ones of the plurality may comprise the same or different detection moieties depending upon the application. Upon exposing the optical elements to an environment where there are analytes of interest, subsequent excitation will generate a luminescent response from those detection moieties that have formed complexes including luminophores.

In this arrangement of bottom excitation, the metal film 250 enhances the excitation intensity via the surface plasmon resonance (SPR) wave generated at the interface by a light source such as a focused laser beam with appropriate polarization. It will be appreciated that to create such SPR that it is desirable that the incident light is capable of creating evanescent waves at the metallic layer dielectric boundary so as to create the necessary generation of those waves in a direction parallel to the interface. The generated SPR can then induce an excitation response from the luminophore component of the complex provided. The system polymer-metal film- spacer-ambient medium selects the range of excitation angles according to its reflection curve. For efficient excitation, as a consequence of a cylindrical symmetry of the optical element illustrated, radial polarization of the excitation beam would be preferred, but other types (unpolarized light, circularly polarized light, linearly polarized light) will work also. It will be appreciated that the nature of the polarization will affect the excitation efficiency achieved.

When the substrate is illuminated from below through the metal film at the plasmon resonance angle the excitation light intensity above the film is strongly enhanced. It will be appreciated that the level of enhancement will depend on the specific s of the structure including for example the thicknesses and nature of the materials used.. This local enhancement can be used to excite a surface bound fluorescence analyte with high efficiency. The proximity of the sample to the metal has also strong influence on the fluorescence emission properties. As was mentioned above with respect to Figures IA, and 1C, the fluorescence emission is highly directional with respect to the polar angle of the substrate surface and the angular distribution of radiation shows a very narrow maximum around the plasmon resonance angle The dielectric space layer 251 between metal 250 and the sample surface is required in order to avoid quenching of the fluorescence which occurs at metal-fluorophore distances below IOnm. This, so called surface-plasmon coupled emission (SPCE) is very surface-specific; thus effectively suppressing fluorescence generated within a bulk of a sample. As a result of the SPCE the generated luminescence is coupled into the optical element within a narrow optical angular range.

One of the key issues is the choice of the spacer layer thickness in order to maximize the detectable fluorescence intensity, i.e., attain a trade-off between SPR excitation and plasmon- coupled intensity. The appropriate choice of the spacer thickness is also required to tune the emission maximum to range of angles collectable by the parabolic element.

Figures 3 and 4 show an alternative configuration for optical elements that may be provided in accordance with the teaching of the present disclosure. In Figure 3, a biochip 300 is provided comprising an array of a plurality of individual optical elements 305, each being formed from a truncated cone structure. Similarly to that described before, the optical elements are upstanding from a common substrate 310 and are typically formed from a plastic material so as to enable fabrication using moulding techniques. A recognition surface or area 315 may be provided on an upper 316 surface of each of the optical elements, the upper surface 316 being separated from the substrate 310 via side walls 317 of the optical element. As shown in the sectional view of Figure 4 a luminescent system incorporating such an array will typically provide a detector 400 below the substrate 310. The detector may be provided in the form of a

CMOS or other suitable detection system. As the generated fluorescence 410 will propagate into the optical element within the narrow light cone resultant from the plasmonic effects- specifϊcally surface plasmon coupled emission, the necessity for an aperture between the substrate and the detector is not essential.

Heretofore the excitation of the detection moiety has been described with reference to an optical excitation from a light source provided below the substrate. Figures 5 to 7 show modifications to such an arrangement where the detection moiety is top illuminated, i.e. the excitation source is provided above the substrate. In this arrangement the excitation source is provided above the optical element. As such it is not possible to generate evanescent waves at the boundary between the metallic layer and the medium where the optical element is used. As a consequence no surface plasmon resonance occurs and, in this way it will be understood that such arrangements will not benefit from surface plasmon resonance enhancement.

However, as the effects of surface plasmon resonance (SPR) and plasmon-coupled emission (SPCE) are independent on each other, such arrangements may still benefit from SPCE, surface plasmon coupled emission. The chip array elements can also be excited from above either individually (Figure 5) or flood-lit by an expanded laser beam (Figures 6 and 7). Although the main advantage of the bottom illumination which was described with reference to the Figures heretofore is lost (enhanced excitation field at the metal-sample interface achieved through SPR), the surface-specificity is believed to be retained as the plasmon coupling occurs only within a certain range of the fiuorophore-interface distance. This type of illumination may be advantageous for certain applications for its simplicity. Furthermore while the configurations are shown in respect of an array of optical elements it will be appreciated that a single optical element modified for plasmonic effects may also be provided in a top lit configuration.

In the arrangement of Figure 5 an array 500 of individual light sources 505, such as may be provided by individual LED's or the like are targeted to as to selectively illuminate individual ones 305 of a plurality of optical elements provided in an array format 300 below. The light 510 emitted from each of the light sources can be targeted or focused onto specific ones of the optical elements below- as shown- or could be targeted onto two or more of the optical elements. By providing such targeted illumination it is possible to discriminate in the excitation provided to each of the detection moieties provided on the individual optical elements.

In the arrangement of Figure 6, the whole top surface of the chip 300 can be illuminated by an expanded beam 600 (i.e. flood-lit) directed downwards. The arrangement of Figure 7 is similar apart from the fact that the light source 700 is offset from the chip, such that the length of the light path travelled from the light source to individual ones of the optical elements varies depending on their relative orientation. This may be usefully employed where a more intense excitation is required at one portion of the chip as opposed to another. Another preferred way to achieve such selective illumination would be through use of filters or the like. Heretofore excitation of the detection moiety has been described with reference to optical excitation. However in a modification to that described, optical elements may be provided which are capable of forming complexes that are responsive to electro-chemi-luminesence excitation. Such complexes will typically incorporate an electro-chemi-luminescence tag or label. Electrochemiluminescence (ECL) is the effect whereby a molecule is caused to be in an excited state by an external electric field-induced chemical reaction. Its depopulation is accompanied by an emission of light in a form of fluorescence or phosphorescence. The major advantage of the ECL-based approach for optical sensing is the low level of the background signal because an optical excitation source is not required. Since the sample is not excited optically, there is no need to suppress the scattered excitation light (e.g. from a laser or LED) or the sample auto- fluorescence. ECL can be usefully combined with SPCE if the plasmon-coupling structure (i.e. metal film) also functions as an electrode for generating the ECL. Such an arrangement may be provided by integrating an ECL dye onto the SPCE layered structure (metal, spacers) of the optical elements. This could then be provided as a single optical element or an array platform featuring high collection efficiency and an inherently low noise level (i.e. a very high signal to noise ratio (SNR) is achievable). Moreover, the use of SPCE here obviates the need for a transparent conducting electrode (such as Indium Tin Oxide) which would typically be required in convention ECL sensor arrangements- the metallic layer that generates the plasmonic effects serving a dual purpose in providing the necessary electrode for the ECL effect. It will be further appreciated that SPCE exhibits a spectral effect whereby the angle of emission depends on the wavelength of the light used for excitation. Using this feature it is possible to provide multiplexed assays on the same optical element by using spectrally distinct ECL labels. Indeed even where the complexes provided do not include ECL labels, an array structure as provided in accordance with the present teaching may be configured to have the response characteristics of a first optical element distinguishable from the response characteristics of a second optical element. By providing optical elements which are configured to associate with complexes of distinguishable spectral emission properties such multiplexed arrays may be provided.

In this way, in order to spatially distinguish the emission peaks, sufficient wavelength separation of the label emission wavelengths is essential. The layered structure also needs to be designed to provide an optimum trade-off between the fluorescence intensities and angular positions and bandwidths for the whole set of labels used

It will be appreciated that what has been described herein are examples of optical elements that utilize plasmonic effects to generate highly efficient detection arrangements. Where provided as an array of optical elements, a chip providing for the parallel and highly efficient detection of fluorescence or other types of luminescence signals is provided. The chip takes advantage of the angular selective propagation of plasmonic effects and large angle optics of for example solid parabolic optical elements and may be used for biodiagnostics including, for example, "lab-on-a-chip" applications. Such an optical chip is designed for the parallel real-time readout of surface-bound fluorescence obtained from biochemical reactions. Fluorescence is obtained from an array of optical elements, each having a receptive molecule optically coupled to its surface. The receptive molecule is capable of detecting the analyte of interest, wherein detection results in luminescence radiated into the optical element. Each optical element in the array may be coated with the same or different detection moiety and is varied by the user based on the analyte of interest.

Figures 8 and 9 show exemplary results that may be achieved using a system incorporating such SPCE configurations. In this experimental result, as shown in Figure 8, an array of nine optical elements 800 (numbered elements 1, 2, 3, 4, 5, 6, 7, 8, & 9) were provided on a shared substrate 810. 50nm Ag and ~25nm PEL (polyelectrolyte multilayer) (PAC- polyacrylic acid final layer) were coated onto all elements to provide the desired metallic and dielectric layers on an upper surface of each of the topical elements. The assembled chip was then incubated for an hour such that a first three of the optical elements (elements 1, 2, 3) were provided with a capture moiety of hlgG (50ug/ml) in PBS (phosphate buffered saline) and the remainder of the optical elements were treated with a blocking agent including BSA (bovine serum albumen) (1% in wt).

A microfluidics arrangement was assembled and the channel flushed with deionised water. The channel was then filled up with Cy5-ahIgG (lug/ml) - in 0.1% PBS-BSA solution. It was subsequently washed with deionised water. As shown in Figure 9, those optical elements that showed an affinity for the Cy5-ahIgG, i.e. formed complexes including luminophores and therefore were capable of providing a luminescent output on excitation exhibited a SPCE optical output providing an intense output light signal in a substantially circular pattern 900. These elements are those identified as elements 2 and 3 from Figure 8, element 1 being contaminated and showing no response. This signal may be easily detected using a suitable positioned detector and shows response characteristics indicative of the presence of the hIgG-Cv5 after 10 minutes. The optical elements that were not capable of forming the complexes having luminophores (elements 4 through 9) show no optical response as a result of exposure to an excitation source.

While not shown in great detail in the present disclosure it will be understood that one or more surfaces of the optical element could be adapted or configured to provide refractive or diffractive optical elements thereon. In this way, light, which would normally impinge on the bottom surface of the substrate at angles greater than the substrate-air critical angle and would thereby undergo total internal reflection, would be transmitted out of the substrate towards the detector.

It will be appreciated that what has been described herein are exemplary arrangements of luminescent sensors configured such that operably plasmonic effects contribute to the output of the sensors. It will be appreciated and understood that these exemplary arrangements are provided to assist in an understanding of the teaching of the present invention but are not to be construed as limiting the scope of the present invention to that described. Where integers or components are described with reference to one figure or example these could be changed for other integers or components and, in this way, it will be understood that various modifications may be made to the embodiments disclosed herein. Therefore, the above description should not be construed as limiting, but merely as exemplification of the various embodiments. Those skilled in the art will envision other modifications within the scope and spirit of the claims appended hereto.

The words comprises/comprising when used in this specification are to specify the presence of stated features, integers, steps or components but does not preclude the presence or addition of one or more other features, integers , steps, components or groups thereof.

Claims

WHAT IS CLAIMED IS:

1. A luminescent sensor chip for operably identifying the presence of one or more analytes within a sensing medium, the chip comprising a substrate with a plurality of optical elements extending upwardly of the substrate, each optical element having a surface configured on exposure to predefined analytes to form a complex on that surface, at least one element of the complex being a luminophore excitation of which operably effects generation of a luminescence signal for subsequent detection, and wherein individual optical elements are configured such that operably plasmonic effects contribute to at least one of the generation or capture of the luminescence signal.

2. The chip of claim 1 wherein the surface is an upper surface of the optical element and has a capture moiety provided thereon, the capture moiety exhibiting a preferential attachment characteristic for predefined analytes.

3. The chip of claim 1 or 2 wherein the surface includes a semi-transparent metallic film and a dielectric spacer layer provided thereon.

4. The chip of any preceding claim wherein on excitation of the luminophores, the resultant detected luminescence signal is enhanced through a combination of at least one of surface plasmon resonance and surface plasmon coupled emission.

5. The chip of claim 3 wherein the optical element is geometrically configured such that on excitation of the luminophores from below, an evanescent wave is formed at a boundary between the optical element and the sensing medium, the evanescent wave effecting generation of surface plasmons.

6. The chip of claim 5 wherein operably the surface plasmons generate surface plasmon resonance which effects enhanced excitation of the luminophores.

7. The chip of claim 4 wherein the surface plasmon coupled emission provides for a coupling of light resultant from the excitation of the luminophores into the optical element at angles greater than or equal to the critical angle of the optical element/sensing medium critical angle.

8. The chip of claim 7 wherein the surface plasmon coupled emission is generated in response to excitation of the luminophores from above or below.

9. The chip of claim 1 wherein each of the optical elements are geometrically configured to totally internally reflect light that enters downwardly into said optical elements from said surface at an angle, θmm, greater than the critical angle of the optical element/sensing medium

10. The chip of claim 9 wherein θmm is about 62 degrees.

11. The chip of claim 3 wherein the semi-transparent metal film and dielectric spacer layer are provided in a layer structure, the dielectric spacer layer operably being the outermost layer adjacent to the sensing medium.

12. The chip of any preceding claim wherein each of the optical elements are substantially paraboloid.

13. The chip of claim 12 wherein the paraboloid elements are solid elements, formed from the same material as the substrate on which they are defined.

14. The chip of any preceding claim wherein the optical elements have side walls and a substantially planar top surface, the top surface at least partially providing the detection surface of the elements.

15. The chip of claim 9 comprising a plurality of annular apertures corresponding to said plurality of optical elements, the apertures being arranged relative to their corresponding optical elements such that light entering said detecting surface at an angle less than Bm_1n impinges on said aperture.

16. The chip of any preceding claim wherein the optical elements are configured such that operably the complexes formed on a first optical element have distinguishable spectral emission properties from the complexes formed on a second optical element.

17. The chip of any preceding claim wherein at least one of the optical elements is configured to operably provide for generation of a complex including an electro-chemi-luminescent tag.

18. A luminescent sensor system including a chip as claimed in any preceding claim, an excitation source configured to operably provide for excitation of the luminophores and a detector configured to provide an output in response to the generated luminescence.

19. The system of claim 18 wherein the excitation source is an optical excitation source.

20. The system of claim 18 wherein the optical excitation source and substrate are arranged relative to one another such that operably the luminophores are excited from below.

21. The system of claim 19 wherein the optical excitation source and substrate are arranged relative to one another such that operably the luminophores are excited from above.

22. The system of claim 19 wherein the excitation source provides for selective illumination of individual ones of the optical elements.

23. The system of claim 22 wherein the excitation source includes a plurality of individual light sources, individual ones of the plurality of lights sources providing optical excitation of specific individual ones of the plurality of optical elements.

24. The system of claim 19 wherein the optical excitation source provides for concurrent illumination of a plurality of optical elements.

25. The system of claim 19 wherein the optical excitation source is arranged so as to be offset from the chip.

26. The system of claim 18 wherein the excitation source is an electro -chemi-luminescent excitation source such that a complex including an electro -chemi-luminescent tag can be caused to be in an excited state by an external electric field-induced chemical reaction.

27. A luminescent sensor system including an optical element having a surface configured on exposure to predefined analytes to form a complex on that surface, at least one element of the complex being a luminophore, excitation of which operably effects generation of a luminescence signal for subsequent detection, the element having a semi-transparent metallic film and a dielectric spacer layer provided thereon such that operably the luminescence signal is coupled into the optical element through surface plasmon coupled emission, the system further including an excitation source configured relative to the optical element to effect excitation of the luminophore from above.

28. The system of claim 27 including a detector provided below the optical element for detection of the generated luminescence signal.

29. The system of claim 27 or claim 28 wherein the surface is configured to form complexes including chemi-luminescent tags such that operably the optical element is responsive to electro- chemi-luminescent excitation.

30. The system of claim 27 wherein the complex is a sandwich assay.