Sensor platform and method for the determination of multiple analytes

• US 20020074513A1
• Filed: 12/04/2001
• Published: 06/20/2002
• Est. Priority Date: 06/05/1999
• Status: Abandoned Application

• Current Assignee: Novartis Ag
• Original Assignee: Novartis Ag
• Inventors: Kresbach, Gerhard Matthias, Duveneck, Gert Ludwig, Abel, Andreas Peter, Ehrat, Markus
• Application Number:
  US10/000,957
• Publication Number:
  US 20020074513A1
• Time in Patent Office: Days
• US Class Current: 250/458.1
• CPC Class Codes: G01N 21/552 Attenuated total reflection
• CPC Class Codes: G01N 21/648 using evanescent coupling or surface plasmon coupling for the excitation of fluorescence
• CPC Class Codes: G01N 21/7703 using reagent-clad optical fibres or optical waveguides using measurement of total internal reflection or attenuated total reflection G01N21/552; optical fibres or waveguides per se G02B
• CPC Class Codes: G01N 21/7743 the reagent-coated grating coupling light in or out of the waveguide

Abstract

The invention is related to a variable embodiment of a sensor platform based on a planar thin-film waveguide for the determination of one or more luminescences from one or more measurement areas on said sensor platform, comprising an optical film waveguide of different layers (“stratified waveguide”) with a first optically transparent layer (a) on a second optically transparent layer (b) of lower refractive index than layer (a) and at least one grating structure for the incoupling of excitation light to the measurement areas or outcoupling of luminescence light from the measurement areas. The invention is also related to an optical system for luminescence determination and to an analytical system, comprising a sensor platform according to the invention, an optical system according to the invention, and supply means for contacting one or more samples with the measurement areas on the sensor platform. Further subjects of the invention are detection methods by luminescence detection, and the use of these methods.

Claims

1. A sensor platform for the simultaneous determination of one or more luminescences from at least two or more, laterally separated measurement areas (d) or at least two or more segments (d′

) comprising several measurement areas, on said platform, comprising an optical film waveguide

with a first optically transparent layer (a) on a second optically transparent layer (b) of lower refractive index than layer (a)

with a grating structure (c) for incoupling excitation light to the measurement areas (d), the grating structure being continuously modulated in the area of the at least two or more measurement areas or of the at least two or more laterally separated segments (d′

) comprising several measurement areas

at least two or more laterally separated measurement areas (d) or at least two or more laterally separated segments (d′

) comprising several measurement areas, and

similar or different biological or biochemical or synthetic recognition elements (e) immobilized in the measurement areas, for the qualitative or quantitative determination of one or more analytes in a sample contacted with the measurement areas,

wherein

the density if the measurement areas on the sensor platform is at least 16 measurement areas per square centimeter, and

a cross-talk of a luminescence, generated in the measurement areas or within a segment and coupled back into the optically transparent layer (a) of the film waveguide, to adjacent measurement areas or adjacent segments is prevented upon outcoupling of this luminescence light by means of the grating structure (c), that is continuously modulated in the area of said measurement areas or segments.

3. A sensor platform according to claim 1 or 2, wherein the grating structure, continuously modulated in the area of the two or more measurement areas or segments, is a superposition of two or more grating structures of different periodicities for the incoupling of excitation light of different wavelenghts, the grating lines being orientated parallel or not parallel, preferably not parallel, to each other, wherein in case of two superimposed grating structures their grating lines are preferably perpendicular to each other.

4. A sensor platform according to any of claims 1-3, wherein an additional optically transparent layer (b′

) with lower refractive index than and in contact with layer (a), and with a thickness of 5 nm-10,000 nm, preferably of 10 nm-1000 nm, is located between the optically between the optically transparent layers (a) and (b).

5. A sensor platform according to any of claims 1-4, wherein an adhesion-promoting layer (f), with a thickness of less than 200 nm, preferably of less than 20 nm, is deposited on the optically transparent layer (a), for immobilization of biological or biochemical or synthetic recognition elements, and wherein the adhesion-promoting layer comprises chemical compounds of the group comprising silanes, epoxides, and “

self-organized functionalized monolayers”

.

6. A sensor platform according to any of claims 1-5, wherein laterally separated measurement areas (d) are generated by laterally selective deposition of biological or biochemical or synthetic recognition elements on the sensor platform, preferably using a method of the group of methods comprising ink jet spotting, mechanical spotting, micro contact printing, fluidic contacting of the measurement areas with the biological or biochemical or synthetic recognition elements upon their supply in parallel or crossed micro channels, upon application of pressure differences or electric or electromagnetic potentials.

7. A sensor platform according to claim 6, wherein, as biological or biochemical or synthetic recognition elements, components of the group comprising nucleic acids (DNA, RNA, . . . ) and nucleic acid analogues (PNA . . . ), antibodies, aptamers, membrane-bound and isolated receptors, their ligands, antigens for antibodies, “

histidin-tag components”

, cavities generated by chemical synthesis, for hosting molecular imprints. etc., are deposited, or wherein whole cells or cell fragments are deposited as biological or biochemical or synthetic recognition elements.

8. A sensor platform according to any of claims 6-7, wherein compounds, preferably of the groups comprising, for example, bovine serum albumin or poly ethylene glycol, which are “

chemically neutral”

towards the analyte, are deposited between the laterally separated measurement areas (d), in order to minimize nonspecific binding or adsorption.

9. A sensor platform according to any of claims 1-8, wherein two or more laterally separated measurement areas are generally combined in segments on the sensor platform, and wherein different segments are preferably additionally separated from each other by a deposited boundary contributing to a fluidic sealing between adjacent areas and/or to a further reduction of optical cross-talk between adjacent segments.

10. A sensor platform according to any of claims 1-9, wherein up to 100,000 measurement areas are provided in a 2-dimensional arrangement and wherein a single measurement area has an area of 0.001 mm²-6 mm².

11. A sensor platform according to any of claims 1-10, wherein the grating structure (c) is a diffractive grating with a uniform period or a multidiffractive grating.

12. A sensor platform according to any of claims 1-10, wherein the grating structure (c) has a laterally varying periodicity in parallel or perpendicular to the direction of propagation of the incoupled light in layer (a).

13. A sensor platform according to any of claims 1-12, wherein the material of the second optically transparent layer (b) comprises quartz, glass, or transparent thermoplastic plastics of the group comprising, for example, poly carbonate, poly imide, or poly methylmethacrylate.

14. A sensor platform according to any of claims 1-13, wherein the refractive index of the first optically transparent layer (a) is higher than 2.

15. A sensor platform according to any of claims 1-13, wherein the first optically transparent layer (a) comprises TiO₂, ZnO, Nb₂O₅, Ta₂O₅, HfO₂, or ZrO₂, especially preferably TiO₂ or Ta₂O₅.

16. A sensor platform according to any of claims 1-15, wherein the thickness of the first optically transparent layer (a) is between 40 and 300 nm, preferably between 70 and 160 nm.

17. A sensor platform according to any of claims 1-16, wherein the grating (c) has a period of 200 nm-1000 nm and a modulation depth of 3 nm-100 nm, preferably of 10 nm-30 nm.

18. A sensor platform according to claims 1 and any of claims 4-16, wherein, by incomplete incoupling and outcoupling of excitation light and/or backcoupled luminescence light, a positive gradient of the intensity of guided excitation light and/or generated luminescence light within a single measurement area and/or across several measurement areas, that can be controlled by means of the grating depth, is generated in parallel to the direction of propagation of the incoupled excitation light.

19. A sensor platform according to claim 18, wherein the grating (c) has a laterally varying grating depth in parallel to the direction of propagation of the incoupled excitation light.

20. A sensor platform according to any of claims 1-17, wherein a negative gradient of the intensity of guided excitation light and/or generated luminescence light within a single measurement area and/or across several measurement areas, that can be controlled by the extent of the propagation losses in the optically transparent layer (a), is generated in parallel to the direction of propagation of the incoupled excitation light.

21. A sensor platform according to claim 17, wherein the ratio of the modulation depth to the thickness of the first optically transparent layer (a) is equal or smaller than 0.2.

22. A sensor platform according to any of claims 1-21, wherein the grating structure (c) is a relief grating with a rectangular, triangular or semi-circular profile or a phase or volume grating with a periodic modulation in the essentially planar, optically transparent layer (a).

23. A sensor platform according to any of claims 1-22, wherein optically or mechanically recognizable marks for simplifying adjustments in an optical system and/or for the connection to sample compartments as part of an analytical system are provided on it.

24. A optical system for the determination of one or more luminescences comprising

at least one excitation light source

a sensor platform according to any of claims 1-23

at least one detector for the collection of the light emanating from one or more measurement areas (d) on the sensor platform.

25. An optical system according to claim 24, wherein the excitation light is launched to the measurement areas in an arrangement of direct or transmission illumination.

26. An optical system according to any of claims 24-25, wherein the detection of the luminescence light is performed in such a way, that the luminescence light outcoupled by a grating structure (c) or (c′

) is collected by the detector as well.

27. An optical system according to claim 24, wherein the excitation light emitted from the at least one light source is coherent and is launched to the one or more measurement areas at the resonance angle for coupling into the optically transparent layer (a).

28. An optical system according to any of claims 24-27, wherein the excitation light from the at least one light source is multiplexed to a plurality of individual rays of as uniform as possible intensity by a diffractive optical element, or in case of multiple lightsources by multiple diffractive optical elements, which are preferably Dammann gratings, or by refractive optical elements, which are preferably microlens arrays, the individual rays being launched essentially parallel to each other to laterally separated measurement areas.

29. An optical system according to any of claims 24-28, wherein two or more coherent light sources of similar or different emission wavelength are used as excitation light sources.

30. An optical system according to claim 29 comprising a sensor platform according to claim 3, wherein the excitation light from two or more coherent light sources is launched simultaneously or sequentially from different directions on a grating structure (c), which comprises a superposition of grating structures of different periodicity.

31. An optical system according to any of claims 24-30, wherein a laterally resolving detector of the group comprising, for example, CCD cameras, CCD chips, photodiode arrays, avalanche diode arrays, multichannel plates and multichannel photomultipliers, is used for signal detection.

32. An optical system according to any of claims 24-31, wherein optical components of the group comprising lenses or lens systems for the shaping of the transmitted light bundles, planar or curved mirrors for the deviation and optionally additional shaping of the light bundles, prisms for the deviation and optionally spectral separation of the light bundles, dichroic mirrors for the spectrally selective deviation of parts of the light bundles, neutral density filters for the regulation of the transmitted light intensity, optical filters or monochromators for the spectrally selective transmission of parts of the light bundles, or polarization selective elements for the selection of discrete polarization directions of the excitation or luminescence light are located between the one or more excitation light sources and the sensor platform according to any of claims 1-23 and/or between said sensor platform and the one or more detectors.

33. An optical system according to any of claims 24-32, wherein the excitation light is launched in pulses with a duration of 1 fsec to 10 min and the emission light from the measurement areas is measured time-resolved.

34. An optical system according to any of claims 24-33, wherein for referencing purposes light signals of the group comprising excitation light at the location of the light sources or after expansion of the excitation light or after its multiplexing into individual beams, scattered light at the excitation wavelength from the location of the one or more laterally separated measurement areas, and light of the excitation wavelength outcoupled by the grating structure (c) besides the measurement areas are measured.

35. An optical system according to claim 34, wherein the measurement areas for determination of the emission light and of the reference signal are identical.

36. An optical system according to any of claims 24-35, wherein launching of the excitation light and detection of the emission light from the one or more measurement areas is performed sequentially for one or more measurement areas.

37. An optical system according to claim 36, wherein sequential excitation and detection is performed using movable optical components of the group comprising mirrors, deviating prisms, and dichroic mirrors.

38. An optical system according to claim 36, wherein sequential excitation and detection is performed using an essentially focus and angle preserving scanner.

39. An optical system according to any of claims 36-38, wherein the sensor platform is moved between steps of sequential excitation and detection.

40. An analytical system for the determination of one or more analytes in at least one sample on one or more measurement areas on a sensor platform by luminescence detection, comprising an optical film waveguide, comprising

a sensor platform according to any of claims 1-23

an optical system according to any of claims 24-39,

supply means for contacting the one or more samples with the measurement areas on the sensor platform.

41. An analytical system according to claim 40, wherein said analytical system additionally comprises one or more sample compartments, which are at least in the area of the one or more measurement areas or of the measurement areas combined to segments open towards the sensor platform, wherein the sample compartments preferably have a volume of 0.1 nl-100 μ

l.

42. An analytical system according to claim 41, wherein the sample compartments are closed, except for inlet and/or outlet openings for the supply or outlet of samples, at their side opposite to the optically transparent layer (a), and wherein the supply or the outlet of the samples and optionally of additional reagents is performed in a closed flow through system, wherein, in case of liquid supply to several measurement areas or segments with common inlet and outlet openings, these openings are preferably addressed row by row or column by column.

43. An analytical system according to claim 41, wherein the sample compartments have openings for locally addressed supply or removal of samples or other reagents at their side opposite to the optically transparent layer (a).

44. An analytical system according to any of claims 41-43, wherein compartments for reagents are provided, which reagents are wetted during the assay for the determination of the one or more analytes and contacted with the measurement areas.

46. A method according to claim 45, wherein the excitation light for the measurement areas is coupled into the optically transparent layer (a) by the grating structure (c).

48. A method for the determination of one or more analytes by luminescence detection on a sensor platform according to any of claims 18-20, wherein the dynamic range for signal measurement and/or quantitative analyte determination can be increased or limited by means of a controllable gradient of guided excitation light and/or excited luminescence light in parallel to the direction of propagation of the incoupled excitation light, within one and/or across several measurement areas.

50. A method according to claim 49, wherein the luminescence label is bound to the analyte or, in a competitive assay, to an analyte analogue or, in a multi-step assay, to one of the binding partners of the immobilized biological or biochemical or synthetic recognition elements or to the biological or biochemical or synthetic recognition elements.

2. A sensor platform for the simultaneous determination of one or more luminescences from at least two or more, laterally separated measurement areas (d) or at least two or more segments (d′

) comprising several measurement areas, on said platform, comprising an optical film waveguide

with a first optically transparent layer (a) on a second optically transparent layer (b) of lower refractive index than layer (a)

with a grating structure (c), that is continuously modulated in the area of the at least two or more-measurement areas or of the at least two or more laterally separated segments (d′

) comprising several measurement areas

at least two or more laterally separated measurement areas (d) or at least two or more laterally separated segments (d′

) comprising several measurement areas, and

similar or different biological or biochemical or synthetic recognition elements (e) immobilized in the measurement areas, for the qualitative or quantitative determination of one or more analytes in a sample contacted with the measurement areas,

wherein

the density of the measurement areas on the sensor platform is at least 16 measurement areas per square centimeter, and

a cross-talk of a luminescence, generated in the measurement areas or within a segment and coupled back into the optically transparent layer (a) of the film waveguide, to adjacent measurement areas or adjacent segments is prevented upon outcoupling of this luminescence light by means of the grating structure (c), that is continuously modulated in the area of said measurement areas.

45. A method for the simultaneous determination by luminescence detection of one or more analytes in one or more samples on at least two or more, laterally separated measurement areas on a sensor platform for the simultaneous determination of one or more luminescences from an array of at least two or more laterally separated measurement areas (d) or at least two or more laterally separated segments (d′

) comprising several measurement areas on said platform, comprising an optical film waveguide

with a first optically transparent layer (a) on a second optically transparent layer (b) of lower refractive index than layer (a)

with a grating structure (c), that is continuously modulated in the area of the at least two or more measurement areas or of the at least two or more laterally separated segments (d′

) comprising several measurement areas

at least two or more laterally separated measurement areas (d) or at least two or more laterally separated segments (d′

) comprising several measurement areas, and

similar or different biological or biochemical or synthetic recognition elements (e) immobilized in the measurement areas, for the qualitative or quantitative determination of one or more analytes in a sample contacted with the measurement areas,

wherein

the density if the measurement areas on the sensor platform is at least 16 measurement areas per square-centimeter, and

a cross-talk of a luminescence, generated in the measurement areas or within a segment and coupled back into the optically transparent layer (a) of the film waveguide, to adjacent measurement areas or adjacent segments is prevented upon outcoupling of this luminescence light by means of the grating structure (c), that is continuously modulated in the area of said measurement areas or segments.

47. A method for the determination of one or more luminescences according to any of claims 45-46, wherein (1) the isotropically emitted luminescence or (2) the luminescence that is coupled back into the optically transparent layer (a) and outcoupled by the grating structure (c) or luminescences of both parts (1) and (2) simultaneously are measured.

49. A method according to any of claims 45-48, wherein for generation of the luminescence a luminescent dye or nanoparticle is used as a luminescence label, which can be excited and emits at a wavelength between 300 nm and 1100 nm.

51. A method according to any of claims 49-50, wherein a second or more luminescence labels of similar or different excitation wavelength as the first luminescence label and similar or different emission wavelength are used.

52. A method according to claim 51, wherein the second or more luminescence labels can be excited at the same wavelength as the first luminescence label, but emit at other wavelengths.

53. A method according to claim 51, wherein the excitation and emission spectra of the applied luminescent dyes do not or only partially overlap.

54. A method according to claim 51, wherein charge or optical energy transfer from a first luminescent dye acting as a donor to a second luminescent dye acting as an acceptor is used for the detection of the analyte.

55. A method according to any of claims 45-54, wherein, besides determination of one or more luminescences, changes of the effective refractive index on the measurement areas are determined.

56. A method according to any of claims 45-55, wherein the one or more luminescences and/or determinations of light signals at the excitation wavelengths are performed polarization-selective, wherein preferrably the one or more luminescences are measured at a polarization that is different from the one of the excitation light.

57. A method according to any of claims 45-56 for the simultaneous or sequential, quantitative or qualitative determination of one or more analytes of the group comprising antibodies or antigens, receptors or ligands, chelators or “

histidin-tag components”

, oligonucleotides, DNA or RNA strands, DNA or RNA analogues, enzymes, enzyme cofactors or inhibitors, lectins and carbohydrates.

58. A method according to any of claims 45-57, wherein the samples to be examined are naturally occurring body fluids, such as blood, serum, plasma, lymph or urine or egg yolk or optically turbid liquids or surface water or soil or plant extracts or bio- or process broths or are taken from biological tissue.

59. The use of a method according to any of claims 45-58 for the determination of chemical, biochemical or biological analytes in screening methods in pharmaceutical research, combinatorial chemistry, clinical and preclinical development, for real-time binding studies and the determination of kinetic parameters in affinity screening and in research, for qualitative and quantitative analyte determinations, especially for DNA- and RNA analytics, for the generation of toxicity studies and the determination of expression profiles and for the determination of antibodies, antigens, pathogens or bacteria in pharmaceutical product development and research, human and veterinary diagnostics, agrochemical product development and research, for patient stratification in pharmaceutical product development and for the therapeutic drug selection, for the determination of pathogens, nocuous agents and germs, especially of salmonella, prions and bacteria, in food and environmental analytics.