Amplified signal in binding assays

• US 7,635,571 B2
• Filed: 12/07/2000
• Issued: 12/22/2009
• Est. Priority Date: 12/07/2000
• Status: Active Grant

• Current Assignee: Siemens Healthcare Diagnostics Products GmbH (Siemens AG)
• Original Assignee: Siemens Healthcare Diagnostics Products GmbH (Siemens AG)
• Inventors: De Keczer, Steve, Davalian, Dariush, Singh, Rajendra, Ullman, Edwin F.
• Primary Examiner(s): BROWN, MELANIE YU
• Application Number:
  US09/732,047
• Publication Number:
  US 20040175696A1
• Time in Patent Office: 3,302 Days
• Field of Search: 435/4, 435/6, 435/7.1, 435/7.4, 435/91.53, 435/91.2, 435/7.5, 435 792- 795, 436/501, 436/518, 436523-534, 436/164, 436/172, 436/905, 536/24.3, 536/24.33, 536/25.32, 977/711
• US Class Current: 435/7.94
• CPC Class Codes: G01N 33/531 Production of immunochemical test materials
• CPC Class Codes: G01N 33/542 with steric inhibition or signal modification, e.g. fluorescent quenching

Abstract

Methods are disclosed for determining minute quantities of an analyte in a medium suspected of containing the analyte. One method comprises treating a medium suspected of containing an analyte under conditions such that the analyte, if present, causes a substrate having an oxidant cleavable linker and a photosensitizer to come into close proximity. The photosensitizer generates singlet oxygen which oxidatively cleaves the linker to form a product which can be detected in a sandwich detection assay such as LOCI. The amount of product detected is related to the amount of analyte in the medium. Compositions, kits, and compounds are also disclosed.

Claims

1. A method for determining the presence or concentration of an analyte in a medium, said method comprising:

providing a reaction mixture comprising in combination;

a medium suspected of containing an analyte;

a first specific binding pair member bound to a water-insoluble solid support;

a second specific binding pair member bound to a sensitizer, said sensitizer capable in its excited state of generating a reactive oxygen species, wherein the proximity of the first specific binding pair member with the second specific binding pair member is modulated by the presence of the analyte; and

digoxigenin-linked biotin linked to the solid support through a reactive oxygen cleavable linker;

incubating the reaction mixture;

exciting the sensitizer, said excitation of the sensitizer causing the formation of reactive oxygen, which cleaves the cleavable linker and releases digoxigenin-linked biotin from the solid support; and

detecting the released digoxigenin-linked biotin, the amount thereof being related to the amount of analyte in said medium.

2. The method of claim 1 wherein:

the proximity of the first and second specific binding pair members to one another results from the binding of the first and second specific binding pair members to the analyte;

the sensitizer is a photosensitizer;

the reactive oxygen species is singlet oxygen; and

the excitation step comprises irradiation of the photosensitizer with light.

3. The method of claim 1 wherein:

the step of detecting the released digoxigenin-linked biotin is carried out by a detection method employing, as a third specific binding pair member, avidin bound to a member of a signal producing system or anti-digoxigenin antibodies bound to a member of a signal producing system or both.