Ultrasensitive immunoassays
First Claim
1. Reagents for use in an assay for detection of a macromolecule comprising two affinity reagents, each reagent being specific for a different determinant of said macromolecule, and each reagent having attached thereto an oligonucleotide which is conjugatable to the oligonucleotide attached to the other of said two reagents, wherein said oligonucleotides are conjugatable by means of ligation or hybridization to each other only when said affinity reagents have became bound to the same said macromolecule and wherein said conjugated oligonucleotides may then serve as a template for a nucleic acid amplification reaction, for detection of said macromolecule.
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Abstract
The present invention relates to an immunological test kit and immunoassay using a first immobilized antibody having affinity for a specific antigen. The invention is characterized by a second and third antibody being specific for different determinants of the antigen and modified with cross-linkable oligonucleotides. For detection, the oligonucleotides are amplified, whereby only such oligonucleotides will be amplified which have been cross-linked to each other. In this way unspecific background is avoided and detection is possible down to single molecules.
48 Citations
15 Claims
- 1. Reagents for use in an assay for detection of a macromolecule comprising two affinity reagents, each reagent being specific for a different determinant of said macromolecule, and each reagent having attached thereto an oligonucleotide which is conjugatable to the oligonucleotide attached to the other of said two reagents, wherein said oligonucleotides are conjugatable by means of ligation or hybridization to each other only when said affinity reagents have became bound to the same said macromolecule and wherein said conjugated oligonucleotides may then serve as a template for a nucleic acid amplification reaction, for detection of said macromolecule.
Specification