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US 2014/0272933 A1Published application
Hcv antigen-antibody combination assay and methods and compositions for use therein
AssigneeAbbott Laboratories
InventorsGeorge J. Dawson, Furesh M. Desai, Robin A. Gutierrez, A. Scott Muerhoff
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Abstract
The present invention generally relates to combination immunoassays, reagents and kits for simultaneous detection of HCV antigens and anti-HCV antibodies in a test sample.
Claims22
An immunoassay for the combined detection of HCV antigen and HCV antibody in a test sample comprising:
- a) simultaneously providing the following reagents:
- i. a solid phase capable of binding to biotin
- ii. biotinylated anti-HCV antibody for the capture of an HCV antigen present said test sample;
- iii. a biotinylated HCV antigen for the capture of anti-HCV antibody in said test sample;
- and
- iv. a detectably labeled HCV antigen for binding to anti-HCV antibody captured by the biotinylated HCV antigen of (iii);
- and
- b) incubating the reagents of step (a) under conditions to produce a reaction mixture that
- (i) the biotinylated anti-HCV antibody of (a)(ii) binds to said solid phase through said biotin and specifically binds to HCV antigen present in said test sample to produce an anti-HCV antibody-HCV antigen complex captured on said solid phase;
- (ii) the biotinylated antigen of (a)(iii) binds to said solid phase through said biotin and specifically binds to anti-HCV antibodies present in said test sample to produce an HCV antigen-anti-HCV antibody complex captured on said solid phase and said detectably labeled HCV antigen of (a)(iv) specifically binds to the anti-HCV antibody in said an HCV antigen-anti-HCV antibody complex captured on said solid phase;
- c) isolating solid phase that comprises attached captured antibody, and captured HCV antigen from unreacted test sample and reagents
- d. contacting the isolated solid phase with a detectably labeled conjugate antibody that binds to said HCV antigen captured in the an anti-HCV antibody-HCV antigen complex of (b)(ii);
- and
- e. detecting the signal generated from the detectable label moieties upon triggering of said signal, wherein presence of said signal indicates presence of HCV in said test sample.
11 more elements
An immunoassay for the simultaneous detection of both HCV antigens and HCV antibodies in a test sample, wherein said combination assay comprises:
- a. a first capture antigen comprising a peptide sequence of a first HCV protein;
- b a first detection antigen comprising a peptide sequence of a first HCV protein and further comprising a detectable label
- c. a second capture antigen comprising an antigenic sequence from a second HCV protein
- d. a second detection antigen comprising an antigenic sequence from a second HCV protein and further comprising a detectable label
- e. a third capture antigen comprising an antigenic sequence from a third HCV protein
- f. a third detection antigen comprising an antigenic sequence from a third HCV protein and further comprising a detectable label
- g. a first capture antibody
- h. a conjugate antibody comprising a detectable label
- wherein said capture antibody and said conjugate antibody specifically bind a fourth HCV protein from said test sample, and said combination assay is performed by:
- (i) contacting said test sample with said capture antigen, said detection antigen, said capture antibody and said conjugate antibody under conditions to allow:
- a) formation of a sandwich complex between said first capture antigen and said detection antigen and first anti-HCV antibody present in said test sample;
- b) formation of a sandwich complex between said second capture antigen and said second detection antigen and an anti-HCV antibody against said second HCV protein present in said test sample;
- c) formation of a sandwich complex between said third capture antigen and said third detection antigen and an anti-HCV antibody against said third HCV protein present in said test sample;
- and
- d) formation of a complex between said capture antibody, said conjugate antibody and an HCV antigen present in said sample;
- and
- (ii) measuring a signal generated from said detectable labels as a result of formation of said complexes, thereby simultaneously detecting HCV antigens and HCV antibodies present in said sample.
14 more elements
An immunoassay for detection of multiple HCV components from a test sample comprising:
- a. providing a biotin-binding solid phase
- b. contacting said solid phase with a mixture that comprises:
- i. biotinylated first capture antigen, biotinylated second capture antigen, biotinylated third capture antigen and biotinylated antibody specific for a fourth HCV antigen;
- and
- ii detectably labeled first, second, and third detection antigens under conditions and time sufficient for
- (1) immune complexes to form between antibodies in said test sample that are independently immunoreactive with and captured by said first, second and third biotinylated antigens, respectively and HCV proteins in said sample that are immunoreactive with said biotinylated antibody, and
- (2) immune complexes to form between said capture antibodies and the respective first, second and third detectably labeled antigens;
- c. isolating solid phase that comprises attached detectably labeled captured antibodies, and captured fourth HCV antigen from unreacted test sample and reagents
- d. contacting the isolated solid phase with a detectably labeled conjugate antibody that binds to said captured fourth HCV antigen;
- and
- e. detecting the signal generated from the detectably labeled moieties upon triggering of said signal, wherein presence of said signal indicates presence of HCV in said test sample.
8 more elements
20-28. (canceled)
A kit for the simultaneous detection of HCV antigens and antibodies in a sample comprising:
- a first pair of capture antigen and detection antigen for detecting a first anti-HCV antibody against a first HCV protein, wherein said detection antigen is detectably labeled
- a second pair of capture antigen and detection antigen for detecting a second anti-HCV antibody against a second HCV protein;
- wherein said detection antigen is detectably labeled
- a third pair of capture antigen and detection antigen for detecting a third anti-HCV antibody against a third HCV protein, wherein said detection antigen is detectably labeled
- a first pair of capture antibody and conjugate antibody for detecting a fourth HCV protein, wherein said conjugate antibody is detectably labeled.
2 more elements
30-38. (canceled)
Dependencies are inferred from the claim text.