QUANTIFICATION OF COXIELLA-BURNETII BY POLYMERASE CHAIN-REACTION (PCR) AND A COLORIMETRIC MICROTITER PLATE HYBRIDIZATION ASSAY (CMHA)

A colorimetric microtiter plate hybridization assay (CMHA) for the qua ntitative determination of Coxiella burnetii DNA after amplification b y externally controlled polymerase chain reaction (PCR) is described. The quantification assay is based on an enzyme linked immunosorbent as say (ELISA) format. Cloned DNA, representing a sequence complementary to an internal part of the diagnostic amplicon, was noncovalently atta ched to the wells of a microtiter plate. Biotinylated PCR product was hybridized to the immobilized capture probe. Bound product was detecte d via streptavidin horseradish peroxidase. The devised nonisotopic tec hnique allows specific, rapid, and convenient quantification of C. bur netii DNA. Additionally, it is compatible with standard laboratory ELI SA equipment, making this assay amenable to automation and permitting processing of large sample numbers.