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The present invention provides novel binding pair compositions of defined and limited stability comprising nucleic acid detection markers useful for the homogeneous, sensitive detection of analytes. Also provided are methods for the sensitive homogenous detection of analytes, particularly analytes o

The present invention provides novel binding pair compositions of defined and limited stability comprising nucleic acid detection markers useful for the homogeneous, sensitive detection of analytes. Also provided are methods for the sensitive homogenous detection of analytes, particularly analytes of clinical relevance. Kits for preparing binding pairs of the invention and for performing the methods of the invention are also provided.

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1. A method for detecting an analyte, comprising: (a) providing an analyte;(b) providing a binding pair comprising: (i) a first binding member comprising a first specificity molecule coupled to a first nucleic acid; and(ii) a second binding member comprising a second specificity molecule coupled to

1. A method for detecting an analyte, comprising: (a) providing an analyte;(b) providing a binding pair comprising: (i) a first binding member comprising a first specificity molecule coupled to a first nucleic acid; and(ii) a second binding member comprising a second specificity molecule coupled to a second nucleic acid, wherein the first and second nucleic acids form a duplex of defined and limited stability;(c) contacting the binding pair with the analyte, thereby forming a complex;(d) dissociating the duplex;(e) allowing the first and second nucleic acids to reassociate, thereby generating a reformed duplex;(f) extending the 3′ ends of the reformed duplex; and(g) detecting the reformed duplex, thereby detecting the analyte. 2. The method of claim 1, wherein step (d) comprises heating the complex to a temperature in excess of the melting temperature of the duplex. 3. The method of claim 1 or 2, wherein the complex is formed in an aqueous solution comprising a salt and step (d) comprises decreasing the salt concentration of the aqueous solution. 4. The method of claim 3, wherein detecting the reformed duplex is accomplished by incorporating a detection molecule into at least one of: the first nucleic acid, the second nucleic acid, and the duplex region of the binding pair. 5. The method of claim 4, wherein the detection molecule is selected from the group consisting of: a fluorescent molecule, a fluorescence quencher molecule, a chemiluminescent compound, a chemiluminescence quencher molecule, a bioluminescent molecule, a radioactive molecule and a fluorescent nucleotide. 6. The method of claim 1, wherein step (d) further comprises diluting the complex. 7. The method of claim 1, wherein step (g) comprises amplification of a nucleic acid molecule comprising the reformed duplex and detecting a product of the amplification. 8. The method of claim 7, wherein the amplification is accomplished by PCR, LCR, SDA, or TMA. 9. The method of claim 7, wherein the amplification is accomplished by PCR using primers that bind only to sites generated by extending the 3 ends of the reformed duplex in step (f). 10. The method of any of claims 7 to 9, wherein the product of the amplification is detected by a method selected from the group consisting of: staining with ethidium bromide, silver staining, autoradiography, dot blotting, slot blotting, and southern blotting. 11. The method of any one of claims 1, 2 and 6-9, wherein at least one of the first and second nucleic acids is a chimeric DNA/RNA molecule and the complex of step (c) is digested with RNAse prior to dissociating the duplex in step (d). 12. A method for the detection of PSA, said method comprising: (a) providing a first binding member comprising a monoclonal antibody directed to a first epitope on prostate specific antigen coupled to a first single-strand nucleic acid and a second binding member comprising a monoclonal antibody directed to a second epitope on prostate specific antigen coupled to a second single-strand nucleic acid, wherein the first single-strand nucleic acid hybridizes to the second single-strand nucleic acid, thereby forming a binding pair that is linked through a nucleic acid duplex;(b) contacting the binding pair with a sample, wherein the sample comprises PSA in a solution, thereby forming a binding pair-PSA complex;(c) heating the binding pair-PSA complex to dissociate nucleic acid duplexes;(d) incubating the binding pair-PSA complex under conditions that permit reassociation of binding members bound to PSA, but does not allow substantial reassociation of excess free binding members in the solution; (e) detecting binding pair duplexes, thereby detecting PSA. 13. The method of claim 12, wherein the first and second monoclonal antibodies together form a sandwich pair of monoclonal antibodies to PSA. 14. The method of any of claim 12 or 13, wherein the sample is a selected from the group consisting of a blood sample, a serum sample, a plasma sample or a tissue sample. 15. The method of claim 12 or 13, wherein the first single-strand nucleic acid comprises SEQ ID NO.:1 and the second single-strand nucleic acid comprises SEQ ID NO.:2 or 3. 16. The method of claim 12 or 13, wherein step (c) is performed at 45° C., and step (d) is performed at room temperature. 17. The method of claim 12 or 13, wherein the detecting step comprises amplifying the duplex region. 18. The method of claim 17, wherein the amplifying comprises the steps of: (i) extending the 3′ ends of the duplex with a DNA polymerase, wherein the extension generates at least one primer binding site; and(ii) performing polymerase chain reaction on the extended duplex using at least one primer complementary to the at least one primer binding site of step (i). 19. A method for the detection of a cell, comprising: (a) providing a sample comprising at least one cell;(b) providing a first binding member comprising an antibody directed to an epitope on the cell coupled to a first single-strand nucleic acid, and a second binding member comprising an antibody directed to an epitope on the cell coupled to a second single-strand nucleic, wherein the first single-strand nucleic acid hybridizes to the second single-strand nucleic acid, thereby forming a binding pair that is linked through a nucleic acid duplex;(c) contacting the binding pair with the sample, thereby forming a binding pair-cell complex;(d) dissociating the nucleic acid duplex;(e) incubating the duplex under conditions that permit reassociation of binding members bound to the cell, but does not allow substantial reassociation of excess free binding members, thereby generating a reformed duplex;(f) detecting a nucleic acid containing the reformed duplex, thereby detecting the cell. 20. The method of claim 19, wherein the cell is selected from the group consisting of: a bacterial cell, an animal cell, a plant cell and a fungal cell. 21. The method of claim 20, wherein the animal cell is a human cell. 22. The method of claim 21, wherein the human cell is diseased. 23. The method of claim 19 or 20, wherein the first nucleic acid comprises SEQ ID NO.:1 and the second first nucleic acid comprises SEQ ID NO.:2 or 3. 24. The method of claim 19 or 20, wherein the first and second antibodies are polyclonal antibodies. 25. The method of claim 24, wherein the first and second antibodies are aliquots of the same polyclonal antibody. 26. The method of claim 19, wherein step (d) comprises heating the duplex. 27. The method of claim 26, wherein the heating is performed at about 45° C. 28. The method of claim 26, wherein the heating is performed at temperature above the melting temperature of the duplex. 29. The method of claim 19, wherein step (e) comprises diluting the complex; and incubating the diluted complex at room temperature. 30. The method of claim 29, wherein the complex is diluted at least 10 fold. 31. The method of claim 29, wherein the complex is diluted at least 100 fold. 32. The method of claim 19, wherein step (f) comprises amplifying a nucleic acid comprising the reformed duplex. 33. The method of claim 32, wherein amplifying comprises the steps of: (i) extending the 3′ ends of the duplex with a polymerase, wherein extending generates a binding site for at least one primer; and(ii) performing polymerase chain reaction on the extended duplex using at least one primer complementary to the binding site generated in step (i). 34. The method of claim 33, wherein the at least one primer comprises SEQ ID NO.:4 or 5. 35. The method of claim 19, wherein the first nucleic acid is coupled to the first antibody through its 3′ terminus and the second nucleic acid is coupled to the second antibody through its 5′ terminus. 36. The method of claim 35, wherein at least one of the first and second nucleic acids is attached to its respective antibody through a spacer. 37. A method for detecting an analyte, the method comprising the steps of: (a) providing a sample comprising an analyte;(b) providing a first binding member comprising an antibody that interacts with the analyte coupled to a first single-strand nucleic acid, wherein the first single-strand nucleic acid comprises a 5′ DNA sequence and a 3′ RNA sequence, wherein the first single-strand nucleic acid is coupled to the first binding member through its 5′ end;(c) providing a second binding member comprising an antibody that interacts with the analyte coupled to a second single-strand nucleic acid, wherein the second single-strand nucleic acid comprises DNA coupled to the second binding member through its 3′ end, wherein the second single-strand nucleic acid hybridizes to the first single-strand nucleic acid, thereby forming a binding pair that is linked through a nucleic acid duplex consisting of regions of both DNA-DNA hybrid and DNA-RNA hybrid;(d) contacting the binding pair with the sample, thereby forming a binding pair-analyte complex;(e) digesting the complex with RNase;(f) dissociating the nucleic acid duplex;(g) incubating the duplex under conditions that permit reassociation of binding members bound to the analyte, but does not allow substantial reassociation of excess free binding members, thereby generating a reformed duplex;(h) extending the 3′ end of the first single-strand nucleic acid in the reformed duplex, which has been exposed as a result of the RNase digestion, wherein the extending generates at least one PCR primer binding site;(i) amplifying a nucleic acid comprising the reformed duplex by PCR using at least one primer that binds to the at least one PCR primer binding site generated in step (h); and(j) detecting the amplified nucleic acid of step (i), thereby detecting the analyte.