PCR-enzyme-linked immunosorbent assay and partial rRNA gene sequencing: a rational approach to identifying mycobacteria.

摘要: A PCR-enzyme-linked immunosorbent assay (ELISA) for amplification and rapid identification of mycobacterial DNA coding 16S rRNA was developed. The PCR selectively targeted amplified part the gene from all mycobacteria while simultaneously labelling one strand product with a 5' fluorescein-labelled primer. identity labelled subsequently determined by hybridization to panel species-specific capture probes, which were immobilized via their biotin ends streptavidin-coated microtiter plate. Specific species probe detected colorimetrically an anti-fluorescein enzyme conjugate. able identify 10 Mycobacterium spp. hybridize (All1) also included. By heminested PCR, sensitive enough detect as little fg DNA, is equivalent approximately three bacilli. directly sputa. specificities probes assessed analysis 60 strains corresponding 18 species. Probes Avi1, Int1, Kan1, Xen1, Che1, For1, Mal1, Ter1, Gor1 specific. Tbc1 cross-hybridized terrae amplicon. Analysis 35 tested blind resulted in 34 being correctly identified. This method could be used early cultures may suitable detection concurrent within clinical specimens.