EP0353937A1
Expired / Lapsed
EP
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Method of assay
Assignee: Applied Research Systems ARS Holding NV
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Filing: 1989-07-25
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Granted: —
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Expires: 2009-07-25
Abstract
Abstract The invention relates to an improved assay technique which provides an enhanced signal-to-noise ratio and enhances sensitivity. Also described are means for carrying out such assays.
Inventors
- 👤 John Worthington Attridge
- 👤 Ian Alexander Flintwood Cottage Shanks
Bibliographic Details
Application Number
—
Jurisdiction
EP
Data Source
google_patents
Last Synchronized
2026-11-09 06:48
CPC Classifications
G; G01; G01N; G01N21/00; G01N21/62; G01N21/63; G01N21/64; G01N21/645; G01N21/648; G01N21/17; G01N21/55; G01N21/552; G01N21/553; G01N33/00; G01N33/48; G01N33/50; G01N33/53; G01N33/543; G01N33/54366; G01N33/54373
Claims Analysis
Total Claims: 20
Claim 1
Independent Claim
205 words
A method of assaying for a ligand in a sample which method comprises the steps of (a) incubating the sample in contact with a specific binding partner for the ligand it is desired to detect, the said specific binding partner being carried on one surface of an optical structure and in the presence of a further reagent X being either a fluorescently or phosphorescently labelled ligand analogue specific for the same specific binding partner or a fluorescently or phosphorescently labelled further specific binding partner for the ligand it is desired to detect, said optical structure comprising a substrate and one or more layers of material interposed between said substrate and the layer of specific binding partner carried on the surface of said optical structure; (b) irradiating another surface of said optical structure at a suitable angle or range of angles to the normal such that resonance occurs within or at the surface of the optical structure, said resonance being surface plasmon resonance or long-range surface plasmon resonance and such that fluorescence or phosphorescence is generated; and (c) analysing the fluorescence or phosphorescence generated in order to determine whether, and if desired the extent to which and/or rate at which, it is altered by complex formation.
Claim 2
Independent Claim
205 words
A method of assaying for a ligand in a sample which method comprises the steps of (a) incubating the sample in contact with a specific binding partner for the ligand it is desired to detect, the said specific binding partner being carried on one surface of an optical structure and in the presence of a further reagent X being either a fluorescently or phosphorescently labelled ligand analogue specific for the same specific binding partner or a fluorescently or phosphorescently labelled further specific binding partner for the ligand it is desired to detect, said optical structure comprising a substrate and one or more layers of material interposed between said substrate and the layer of specific binding partner carried on the surface of said optical structure; (b) irradiating another surface of said optical structure at a suitable angle or range of angles to the normal such that resonance occurs within or at the surface of the optical structure, said resonance being surface plasmon resonance or long-range surface plasmon resonance and such that fluorescence or phosphorescence is generated; and (c) analysing the fluorescence or phosphorescence generated in order to determine whether, and if desired the extent to which and/or rate at which, it is altered by complex formation.
Claim 3
Dependent Claim
17 words
A method as claimed in claim 1 wherein, in step (b), said resonance is surface plasmon resonance.
Claim 4
Dependent Claim
18 words
A method as claimed in claim 2 wherein, in step (c), the property analysed is the generated fluorescence.
Claim 5
Independent Claim
162 words
A method of assaying for a ligand in a sample which method comprises the steps of (a) incubating the sample in contact with a specific binding partner for the ligand it is desired to detect, the said specific binding partner being carried on one surface of an optical structure, said optical structure comprising a substrate and one or more layers of material interposed between said substrate and the said specific binding partner; (b) irradiating another surface of said optical structure such that the radiation is totally internally reflected and resonance occurs within the optical structure, said resonance being either long-range surface plasmon resonance or resonant guided mode coupling; and (c) analysing the reflected radiation in order to determine whether, and if desired the extent to which and/or rate at which, the optical characteristics of the sensor formed by the optical structure and specific binding partner carried thereon are altered by formation of a complex between the ligand and the specific binding partner.
Claim 6
Independent Claim
162 words
A method of assaying for a ligand in a sample which method comprises the steps of (a) incubating the sample in contact with a specific binding partner for the ligand it is desired to detect, the said specific binding partner being carried on one surface of an optical structure, said optical structure comprising a substrate and one or more layers of material interposed between said substrate and the said specific binding partner; (b) irradiating another surface of said optical structure such that the radiation is totally internally reflected and resonance occurs within the optical structure, said resonance being either long-range surface plasmon resonance or resonant guided mode coupling; and (c) analysing the reflected radiation in order to determine whether, and if desired the extent to which and/or rate at which, the optical characteristics of the sensor formed by the optical structure and specific binding partner carried thereon are altered by formation of a complex between the ligand and the specific binding partner.
Claim 7
Independent Claim
219 words
A method of assaying for a ligand in a sample which method comprises the steps of (a) incubating the sample in contact with a specific binding partner for the ligand it is desired to detect, the said specific binding partner being carried on one surface of an optical structure and in the presence of a further reagent X being either a fluorescently or phosphorescently labelled ligand analogue specific for the same specific binding partner or a fluorescently or phosphorescently labelled further specific binding partner for the ligand it is desired to detect, said optical structure comprising a substrate and optionally one or more layers of material interposed between said substrate and the said specific binding partner; (b) irradiating another surface of said optical structure in a plane perpendicular to the plane of the said layers of material and at a suitable angle or range of angles to the normal such that the radiation is totally internally reflected and such that fluorescence or phosphorescence is generated; and (c) monitoring the generated fluorescence or phosphorescence which emerges from an edge of said optical structure not in the path of the applied radiation and analysing said fluorescence or phosphorescence in order to determine whether, and if desired the extent to which and or rate at which, it is altered by complex formation.
Claim 8
Independent Claim
219 words
A method of assaying for a ligand in a sample which method comprises the steps of (a) incubating the sample in contact with a specific binding partner for the ligand it is desired to detect, the said specific binding partner being carried on one surface of an optical structure and in the presence of a further reagent X being either a fluorescently or phosphorescently labelled ligand analogue specific for the same specific binding partner or a fluorescently or phosphorescently labelled further specific binding partner for the ligand it is desired to detect, said optical structure comprising a substrate and optionally one or more layers of material interposed between said substrate and the said specific binding partner; (b) irradiating another surface of said optical structure in a plane perpendicular to the plane of the said layers of material and at a suitable angle or range of angles to the normal such that the radiation is totally internally reflected and such that fluorescence or phosphorescence is generated; and (c) monitoring the generated fluorescence or phosphorescence which emerges from an edge of said optical structure not in the path of the applied radiation and analysing said fluorescence or phosphorescence in order to determine whether, and if desired the extent to which and or rate at which, it is altered by complex formation.
Claim 9
Dependent Claim
89 words
A sensor for detecting a ligand in a sample by the method of claim 1 or claim 4 which comprises an optical structure having a substrate, a layer of metal and interposed therebetween a layer of dielectric material having a refractive index lower than that of said substrate, and a specific binding partner for the ligand it is desired to detect adsorbed on or bound to (directly or indirectly) the said metal layer, said layers being such that, in use, long range surface plasmon resonance may be propagated therein.
Claim 10
Dependent Claim
41 words
A sensor as claimed in claim 6 wherein the layer of metal is of silver and is between 10 and 50 nm thick, and the layer of dielectric material is of magnesium fluoride and is between 10 and 2000 nm thick.
Claim 11
Dependent Claim
24 words
A sensor as claimed in claim 7 wherein the silver layer is 15.5 nm thick and the magnesium fluoride layer is 1500 nm thick.
Claim 12
Dependent Claim
81 words
A sensor for detecting a ligand in a sample by the method of claim 4 which comprises an optical structure having a substrate coated with a thin layer of metal, which metal layer is itself coated with a layer of dielectric material of a thickness suitable to support one or more guided modes of radiation of wavelength employed when the sensor is in use and which dielectric layer carries a specific binding partner for the ligand it is desired to detect.
Claim 13
Dependent Claim
31 words
A sensor as claimed in claim 9 wherein the layer of metal is of silver or gold and is about 50nm thick and the layer of dielectric material is of silica.
Claim 14
Dependent Claim
131 words
Apparatus suitable for use in a method of assay as claimed in any one of claims 1 to 5 which comprises (a) a sensor, the said sensor comprising a specific binding partner for a ligand it is desired to assay carried on the surface of an optical structure comprising a substrate and one or more layers of material interposed between said substrate and said specific binding partner; (b) a collimated source of radiation which is capable of being arranged such that, in use, the radiation enters said optical structure at an angle suitable to produce total internal reflection and optionally resonance (said resonance being surface plasmon resonance, long-range surface plasmon resonance or resonant guided mode coupling) within said optical structure; and (c) means for in use analysing reflected or generated radiation.
Claim 15
Dependent Claim
13 words
Apparatus as claimed in claim 11 wherein the source of radiation is polarised.
Claim 16
Dependent Claim
23 words
Apparatus as claimed in claim 11 or claim 12 wherein the sensor is as defined in any one of claims 6 to 10.
Claim 17
Dependent Claim
56 words
A specifically reactive sample-collecting and testing device possessing a cavity or cavities each having a dimension small enough to enable sample liquid to be drawn into the cavity by capillary action, and wherein at least one part of a wall of said cavity comprises a sensor as claimed in any one of claims 6 to 10.
Claim 18
Dependent Claim
112 words
A specifically reactive sample-collecting and testing device possessing a cavity or cavities each having a dimension small enough to enable sample liquid to be drawn into the cavity by capillary action, and wherein at least one part of a wall of said cavity comprises a sensor for detecting a ligand in said sample which comprises a specific binding partner for the ligand it is desired to detect carried on the surface of an optical structure comprising a substrate and, interposed between said specific binding partner and said substrate, a thin layer of metal and optionally a layer of dielectric material such that said optical structure is capable of supporting a surface plasmon.
Claim 19
Dependent Claim
112 words
A specifically reactive sample-collecting and testing device possessing a cavity or cavities each having a dimension small enough to enable sample liquid to be drawn into the cavity by capillary action, and wherein at least one part of a wall of said cavity comprises a sensor for detecting a ligand in said sample which comprises a specific binding partner for the ligand it is desired to detect carried on the surface of an optical structure comprising a substrate and, interposed between said specific binding partner and said substrate, a thin layer of metal and optionally a layer of dielectric material such that said optical structure is capable of supporting a surface plasmon.
Claim 20
Dependent Claim
43 words
A device as claimed in claim 14 or claim 15 wherein the wall of the capillary cavity which is remote from the wall comprising a sensor carries in dry releasable form a fluorescently or phosphorescently labelled ligand analogue or further specific binding partner.
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