EP0100619A1
Expired / Lapsed
EP
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Enzyme chromatographic immunoassay
Assignee: Siemens Healthcare Diagnostics GmbH Germany
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Filing: 1983-07-14
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Granted: —
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Expires: 2003-07-14
Abstract
Abstract Chromatographic immunoassay employing a specific binding pair member and a label conjugate which delineates a border whose distance from one end of the chromatograph relates to the amount of analyte present. By combining the label conjugate and sample in a solution and immunochro- matographing the solution, or employing a combination of enzymes, one enzyme being the label and the other enzyme affixed to the chromatographic support, the position of the border defined by the label can be related to the amount of analyte in the sample solution. Preferably, an immunochromatograph is employed having both a specific binding pair member and an enzyme affixed to the support. A sample is chromatographed and the amount of analyte is determined by (1) contacting the chromatograph with a second enzyme conjugated with a specific binding pair member which binds to the chromatograph in proportion to the amount of analyte bound to the chromatograph, or (2) including the second enzyme conjugate with said sample, resulting in a defined border related to the amount of analyte in the sample. The two enzymes are related in that the substrate of one is the product of the other, so that upon contact of the chromatograph with appopriate reagents, a detectable signal develops which permits detection of the border to which the analyte traveled. This distance can be related to the amount of analyte present in the sample.
Inventors
- 👤 Robert F. Zuk
- 👤 David J. Litman
Bibliographic Details
Application Number
—
Jurisdiction
EP
Data Source
google_patents
Last Synchronized
2026-10-28 21:40
CPC Classifications
G; G01; G01N; G01N33/00; G01N33/48; G01N33/50; G01N33/53; G01N33/543; G01N33/54366; G01N33/54386; G01N33/54387; G01N33/54388; Y; Y10; Y10S; Y10S435/00; Y10S435/805; Y10S435/97; Y10S436/00; Y10S436/807; Y10S436/81; Y10S436/815
Claims Analysis
Total Claims: 15
Claim 1
Independent Claim
304 words
A method for determining the presence of an analyte in a sample suspected of containing said analyte, said method employing: (A) an immunochromatograph having a porous support permitting solvent travel and a plurality of a specific binding pair member ("mip") non-diffusively and uniformly bound to said support and extending a distance away from a first end of said support to define an immunosorbing zone; and (B) a labeled mip, where said labeled mip is chosen to bind to said immunosorbing zone in relation to the portion of said immunosorbing zone in which said analyte is bound to define a border related to the amount of analyte present in said sample, said label being an enzyme or coenzyme which is part of a signal producing system including at least one enzyme which system produces a detectable signal of electromagnetic radiation defining said border; said method comprising: contacting said immunochromatograph: (1) at said first end with sample containing solvent for a time sufficient for at least a portion of said solvent to traverse said immunosorbing zone; and (2) with labeled mip containing solvent, where said labeled mip binds in said immunosorbing zone in relation to the analyte bound in said immunosorbing zone; and determining the border defined by said labeled mip by means of said signal producing system, by contacting said immunosorbing zone with the remaining members of said signal producing system, where the position of the border relates to the amount of anal y te present in said sample; with the proviso that when said immunosorbing zone is contacted stepwise, first with said sample containing solvent, and then with said labeled mip containing solvent, a second enzyme is non-diffusively and uniformly bound in said immunosorbing zone, where the two enzymes are related by the product of one enzyme being the substrate of the other enzyme.
Claim 2
Independent Claim
304 words
A method for determining the presence of an analyte in a sample suspected of containing said analyte, said method employing: (A) an immunochromatograph having a porous support permitting solvent travel and a plurality of a specific binding pair member ("mip") non-diffusively and uniformly bound to said support and extending a distance away from a first end of said support to define an immunosorbing zone; and (B) a labeled mip, where said labeled mip is chosen to bind to said immunosorbing zone in relation to the portion of said immunosorbing zone in which said analyte is bound to define a border related to the amount of analyte present in said sample, said label being an enzyme or coenzyme which is part of a signal producing system including at least one enzyme which system produces a detectable signal of electromagnetic radiation defining said border; said method comprising: contacting said immunochromatograph: (1) at said first end with sample containing solvent for a time sufficient for at least a portion of said solvent to traverse said immunosorbing zone; and (2) with labeled mip containing solvent, where said labeled mip binds in said immunosorbing zone in relation to the analyte bound in said immunosorbing zone; and determining the border defined by said labeled mip by means of said signal producing system, by contacting said immunosorbing zone with the remaining members of said signal producing system, where the position of the border relates to the amount of anal y te present in said sample; with the proviso that when said immunosorbing zone is contacted stepwise, first with said sample containing solvent, and then with said labeled mip containing solvent, a second enzyme is non-diffusively and uniformly bound in said immunosorbing zone, where the two enzymes are related by the product of one enzyme being the substrate of the other enzyme.
Claim 3
Dependent Claim
21 words
A method according to Claim 1, wherein said label is an enzyme and said second enzyme is bound to said immunochromatograph.
Claim 4
Dependent Claim
14 words
A method according to Claim 2, wherein at least one enzyme is an oxidoreductase.
Claim 5
Independent Claim
24 words
A method according to any of Claims 1, 2 or 3, where said labeled mip and said sample are included in the same solvent.
Claim 6
Dependent Claim
39 words
A method according to Claim 4, wherein said labeled mip traverses said immunosorbing zone at a slower rate than said analyte, so as to concentrate at about a border between an analyte containing region and an analyte free region.
Claim 7
Dependent Claim
37 words
A method according to Claim 4, where said determining includes the step of immersing said immunosorbing zone in a solution containing an enzyme substrate which is enzymatically transformed to a product capable of detection by electromagnetic radiation.
Claim 8
Dependent Claim
15 words
A method according to Claim 6, wherein said product absorbs light in the visible range.
Claim 9
Independent Claim
275 words
A method for determining the presence of an analyte in a sample suspected of containing said analyte, said method employing: (A) an immunochromatograph having a porous support permitting solvent travel and non-diffusively and uniformly bound to said porous support a plurality of a specific binding pair member ("mip") and a plurality of a second enzyme, extending a distance away from the first end of said support to define an immunosorbing zone; and (B) a first enzyme labeled mip, where said first and second enzymes are related by the product of one enzyme being the substrate of the other enzyme and said labeled mip is chosen to bind to said immunosorbing zone in relation to the portion of said immunosorbing zone in which said analyte is bound to define a border related to the amount of analyte, and where said enzymes and a substrate which produces a dye which binds to said support are members of a signal producing system; said method comprising: contacting said immunochromatograph at said first end with a solution containing sample and first enzyme labeled mip for a time sufficient for at least a portion of said solution to traverse said immunosorbing zone, where said enzyme labeled mip binds in said immunosorbing zone in relation to the analyte bound in said immunosorbing zone; and contacting said immunosorbing zone in a substrate containing solution resulting in depositing of said dye in the environs of said second enzyme on said support defining a region in which analyte is bound, wherein the distance the border of said region is from one end of said immunochromatograph is related to the amount of analyte in said sample.
Claim 10
Independent Claim
275 words
A method for determining the presence of an analyte in a sample suspected of containing said analyte, said method employing: (A) an immunochromatograph having a porous support permitting solvent travel and non-diffusively and uniformly bound to said porous support a plurality of a specific binding pair member ("mip") and a plurality of a second enzyme, extending a distance away from the first end of said support to define an immunosorbing zone; and (B) a first enzyme labeled mip, where said first and second enzymes are related by the product of one enzyme being the substrate of the other enzyme and said labeled mip is chosen to bind to said immunosorbing zone in relation to the portion of said immunosorbing zone in which said analyte is bound to define a border related to the amount of analyte, and where said enzymes and a substrate which produces a dye which binds to said support are members of a signal producing system; said method comprising: contacting said immunochromatograph at said first end with a solution containing sample and first enzyme labeled mip for a time sufficient for at least a portion of said solution to traverse said immunosorbing zone, where said enzyme labeled mip binds in said immunosorbing zone in relation to the analyte bound in said immunosorbing zone; and contacting said immunosorbing zone in a substrate containing solution resulting in depositing of said dye in the environs of said second enzyme on said support defining a region in which analyte is bound, wherein the distance the border of said region is from one end of said immunochromatograph is related to the amount of analyte in said sample.
Claim 11
Dependent Claim
14 words
A method according to Claim 8, wherein said first and second enzymes are oxidoreductases.
Claim 12
Dependent Claim
20 words
A method according to Claim 9, wherein said first enzyme is a peroxidase and said second enzyme produces hydrogen peroxide.
Claim 13
Dependent Claim
14 words
A method according to Claim 10, wherein said second enzyme is an oxidase. ,
Claim 14
Dependent Claim
15 words
A method according to Claim 8, wherein said dye is deposited primarily at said border.
Claim 15
Dependent Claim
19 words
A method according to Claim 8, wherein said dye is deposited throughout said region in which analyte is bound.
References Cited (Backward Citations)
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