A SENSITIVITY MULTIANALYTE IMMUNOASSAY USING COVALENT DNA-LABELED ANTIBODIES AND POLYMERASE CHAIN-REACTION

A multianalyte immunoassay for simultaneous detection of three analyte s (hTSH, hCG and beta-Gal) has been demonstrated using DNA-labeled ant ibodies and polymerase chain reaction (PCR) for amplification of assay response. The labeled antibodies were prepared by covalently coupling uniquely designed DNA oligonucleotides to each of the analyte-specifi c monoclonal antibodies. Each of the DNA oligonucleotide labels contai ned the same primer sequences to facilitate coamplification by a singl e primer pair. Assays were performed using a two-antibody sandwich ass ay format and a mixture of the three DNA-labeled antibodies. Dose-resp onse relationships for each analyte were demonstrated. Analytes were d etected at sensitivities exceeding those of conventional enzyme immuno assays by approximately three orders of magnitude. Detection limits fo r hTSH, beta-Gal and hCG were respectively 1 x 10(-19), 1 x 10(-17) an d 1 x 10(-17) mol. Given the enormous amplification afforded by PCR an d the existing capability to differentiate DNA based on size or sequen ce differences, the use of DNA-labeled antibodies could provide the ba sis for the simultaneous detection of many analytes at sensitivities g reater than those of existing antigen detection systems. These finding s in concert with previous reports suggest this hybrid technology coul d provide a new generation of ultra-sensitive multianalyte immunoassay s.