Simultaneous calibration heterogeneous immunoassay

• US 4,843,000 A
• Filed: 05/21/1985
• Issued: 06/27/1989
• Est. Priority Date: 12/26/1979
• Status: Expired due to Term

• Current Assignee: Dade Behring Marburg GmbH (Siemens AG)
• Original Assignee: Syntec Incorporated (Atos SE)
• Inventors: Ullman, Edwin F., Litman, David J.
• Primary Examiner(s): Kepplinger, Esther M.
• Application Number:
  US06/736,493
• Time in Patent Office: 1,498 Days
• Field of Search: 435/4, 435/7, 435/188, 435/805, 435/810, 436/524, 436/525, 436/527-529, 436/531, 436/546, 436/808, 436/810, 436/824, 422/55, 422/56, 422/61
• US Class Current: 435/7.91
• CPC Class Codes: G01N 33/52 Use of compounds or compositions for colorimetric, spectrophotometric or fluorometric investigation, e.g. use of reagent paper and including single- and multilayer analytical elements immunological elements G01N33/54386; involving labelled immunochemicals G01N33/58; for haemoglobin or occult blood G01N33/72
• CPC Class Codes: G01N 33/542 with steric inhibition or signal modification, e.g. fluorescent quenching
• CPC Class Codes: G01N 33/543 with an insoluble carrier for immobilising immunochemicals
• CPC Class Codes: G01N 33/54306 Solid-phase reaction mechanisms
• CPC Class Codes: G01N 33/581 with enzyme label (including co-enzymes, co-factors, enzyme inhibitors or substrates)
• CPC Class Codes: Y10S 435/805 Test papers
• CPC Class Codes: Y10S 436/81 Tube, bottle, or dipstick

Abstract

An assay method and compositions are provided for determining the presence of an analyte in a sample. The analyte is a member of an immunological pair (mip) comprising ligand and receptor. By providing a first measurement surface capable of specifically binding a labelled reagent in an amount depending upon the presence of analyte in the sample and a second calibration surface capable of binding a second labeled reagent in a manner unaffected by the presence of analyte in the sample, calibration of individual tests can be accomplished simulataneously with the performance of the test itself. A signal producing system includes an enzyme bonded to a mip which defines the first labeled reagent for binding to the measurement surface and the same enzyme conjugated to a ligand capable of binding to the calibration surface. Preferably, both labeled reagents have the same composition and the calibration surface includes anti-(first enzyme).

Claims

1. In a method for determining the presence in a sample of an analyte, which analyte is a member of a specific binding pair ("mip") consisting of ligand and receptor ("antiligand"),

said method employing at least one catalyst including a catalyst bound to a mip ("catalyst-bound-mip") and a solute which is catalytically transformed by a catalyst bound to a mip-containing measurement first surface to produce a detectable signal at said first surface in proportion to the amount of catalyst-bound-mip bound to said first surface, wherein contacting of said first surface with said sample and said catalyst-bound-mip results in binding of said catalyst-bound-mip to said first surface in proportion to the amount of analyte in said sample,

the improvement which comprises:

having adjacent to said first surface a calibration second surface to which said catalyst is directly bound in an amount to produce a signal level at said second surface corresponding to a predetermined amount of analyte and said analyte in said sample is determined by comparing the intensity of the signal at said first surface to the intensity of the signal at said second surface.

2. A method according to claim 1, wherein said catalyst is an enzyme.

4. A method according to any of claims 1, 2, or 3, wherein said signal producing system includes two enzymes as catalysts, wherein the product of one enzyme is the substrate of the other enzyme, and one of the enzymes is bound to each of said first and second surfaces prior to said contacting.

5. A method according to claim 4, wherein said solute is a dye precursor which undergoes an enzyme catalyzed reaction to produce an insoluable dye which binds to said surface.

6. A method according to claim 5, wherein said catalyst-bound-mip is enzyme-bound-antiligand.

7. A method according to claim 5, wherein said catalyst-bound-mip is enzyme-bound-ligand.

3. In a method for determining the presence in a sample of an analyte, which analyte is a member of a specific binding pair ("mip") consisting of ligand and receptor ("antiligand"),

said method employing at least one catalyst including catalyst bound to a mip ("catalyst-bound-mip") and a solute which is catalytically transformed by a catalyst bound to a mip-containing measurement first surface to produce a detectable signal at said first surface in proportion to the amount of catalyst-bound-mip bound to said first surface, wherein contacting of said first surface with said sample and said catalyst-bound-mip results in binding of said catalyst-bound-mip to said first surface in proportion to the amount of analyte in said sample,

the improvement which comprises:

having adjacent to said first surface a calibration second surface to which said first catalyst binds by means of mip complex formation in an amount to produce a signal level at said second surface corresponding to a pre-determined amount of analyte, wherein the binding of said catalyst to said calibration second surface involves a determinant site on said catalyst or a determinant site on the catalyst-bound-mip other than the determinant site involved in the binding of said catalyst-bound-mip to said measurement first surface, whereby said analyte in said sample is determined by comparing the intensity of signal at said second surface to the intensity of signal at said first surface.

8. In a method for determining the presence in a sample of an analyte, which analyte is a member of a specific binding pair ("mip") consisting of ligand and receptor "antiligand")

said method employing a signal producing system having at least two enzymes, including one enzyme bound to a mip ("enzyme-bound-mip"), and a solute dye precursor which is catalytically transformed to an insoluable dye by one of said enzymes bound to a mip-containing measurement first surface, said insoluble dye producing a detectable signal at said first surface in proportion to the amount of analyte, where said mip at said first surface provides for binding of enzyme-bound-mip through mip complex formation to said first surface, wherein contacting of said first surface with said sample and said enzyme-bound-mip results in binding of said enzyme-bound-mip to said surface in proportion to the amount of analyte in said sample,

the improvement which comprises:

during said contacting, having adjacent to said first surface a calibration second surface to which enzyme of said enzyme-bound-mip becomes bound through mip complex formation in an amount to produce a signal level at said second surface corresponding to a pre-determined amount of analyte, wherein the mip complex formation on said calibration second surface involves a determinant site on said enzyme or a dterminant site on the enzyme-bound-mip other than the determinant site involved in the mip complex formation on said measurement first surface, whereby said analyte in said sample is determined by comparing the intensity of signal at said second surface to the intensity of signal at said first surface.