RAPID DETECTION OF ENTEROVIRUS IN CLINICAL SPECIMENS USING PCR AND MICROWELL CAPTURE HYBRIDIZATION ASSAY

A rapid detection method of enteroviral RNA in clinical samples using PCR and a microwell capture hybridization assay is described. PCR prod ucts were labelled directly by digoxigenin-dUTP during the amplificati on step. The labelled amplicons were hybridized with a biotinylated ol igo-probe and captured on commercially available test microwells coate d with streptavidin. The hybridized amplicons labelled with digoxigeni n were detected using anti-digoxigenin Fab fragments conjugated to per oxidase and colorimetric reaction automatically measured. This method detected as few as 0.01 PFU/100 mu l of biological sample with a resul t obtained within 8 h. Using this method, we were able to detect enter oviral RNA in 23 of 35 clinical specimens from 16 of 17 patients with suspected acute or chronic enteroviral infection. The samples included cerebrospinal fluid, broncho-pulmonary lavage, pericardial effusion, throat swabs, stools, sera, muscular and myocardial biopsies. In contr ast, virus was isolated in cell culture in only 8 of 28 clinical speci mens from 6 of the 17 patients. This easy-to-perform assay has useful potential in the rapid detection of enterovirus in acute or chronic in fection. This methodology could be used for a rapid qualitative detect ion of other RNA viruses