QUANTITATIVE DETECTION OF HEPATITIS-C VIRUS-RNA BY MULTICYCLIC RT-PCR

We have developed a method to quantitate hepatitis C virus (HCV)-RNA from serum by using 'multicylic' RT-PCR. This method requires 100 mul of sera and synthetic HCV-RNA with a known copy number to be used as a standard. HCV-RNA in serum are amplified by the RT-PCR technique and quantitated by comparing the darkness of the spot on the film with the standards after dot blot hybridization. Accuracy of this technique was comparable to the competitive RT PCR method (CRT-PCR) when compared using the same sample. Also, this technique uses one-tenth of the sample volume required in CRT-PCR with the additional advantages of low cost, faster assay time, and ability to run a larger number of samples. We analyzed the amount of HCV-RNA in 71 patients with chronic liver disease due to HCV subclassified into chronic inactive hepatitis (CIH), chronic active hepatitis (CAH), liver cirrhosis (LC), and hepatocellular carcinoma (HCC). Seventy-five percent of the patients contained 10(6) to 10(8) copies/ml of HCV-RNA. The number of samples which contained over 10(6) copies/ml in the four groups were CIH, 5/8 (62.5%); CAH, 15/20 (75.0%); LC, 25/27, (92.6%); and HCC, 14/16, (87.5%).