Microtitre plate riboprobe system for detection of ultrasonicated Campylobacter jejuni genomic DNA.

ABSTRACT

Microtitre plate nucleic acid probe hybridization systems were developed for the detection of thermophilic Campylobacter and Campylobacter jejuni. Specific RNA probes obtained by in vitro transcription of DNA templates synthesized by polymerase chain reaction using two sets of specific primers incorporating bacteriophage T7 promoter sequences were immobilized on a microtitre plate. The hybridizations were carried out on samples of genomic DNA sheared by ultrasonication. Optimum conditions for the ultrasonic treatment were determined in order to obtain the highest degree of hybridization with immobilized RNA probe. Finally, detection of RNA-DNA hybrids in the wells was accomplished by an immunoenzymatic assay using a monoclonal anti-RNA-DNA hybrid antibody. This rapid, simple hybridization and immunoenzymatic assay system will facilitate the detection of Campylobacter in foods and clinical samples.