AUTOMATED QUANTITATIVE-DETERMINATION OF HEPATITIS-C VIRUS VIREMIA BY REVERSE TRANSCRIPTION PCR

An automated reverse transcription PCR was developed for the quantitat ive detection of hepatitis C virus. The quantitation is based on the c oamplification and labelling with dig xigenin-dUTP during PCR of two s imilar templates, the viral genome and a modified RNA which acts as a mimic target. Known amounts of the mimic RNA sequence were introduced into the clinical samples. The automated quantitation of the two coamp lified and labelled products depends on the use of two biotinylated ca pture probes which are complementary, respectively, to a deleted seque nce and to an inserted sequence introduced by site-directed mutagenesi s in a wild viral cloned cDNA. This method proved to be simple, reprod ucible, and useful for quantitate hepatitis C virus viremia in chronic ally infected patients. This easy-to-perform, automated assay could al so be used for the accurate determination of human immunodeficiency vi rus viremia or other RNA molecules.