A QUANTITATIVE HCV-PCR TEST FOR ROUTINE DIAGNOSTICS

The aim of this study was to develop a reliable and simple method for hepatitis C virus (HCV)-PCR using standard, automated laboratory equip ment. HCV-RNA was extracted from serum and amplified in a single PCR w ith an internal standard. The PCR product was detected using fluoroimm unoassay. Quantification was based on external and internal standards. Linearity was observed over a wide range (10 - 10(7) geq). Mean inter and intra serial coefficients of variation were 35% and 23%, respecti vely. The limit of quantification was 1000 geq/ml based on intra and i nter serial variations, while levels of 110 geq/ml were always detecta ble. Lower concentrations were intermittently positive. The ability to separate HCV-signals in healthy and infected persons was good, based on the distribution of HCV-signals from 353 random blood donors and 19 1 patient samples. To illustrate the applicability of the test, HCV-RN A quantification was performed in 11 patients during treatment with in terferon alpha-2b. Ten of 11 patients showed a decline in HCV-RNA with in the first few weeks of treatment. After four weeks most patients we re still HCV-RNA positive but below the limit of quantification. The p resent method for quantification of HCV-RNA was shown to have sensitiv ity at the level of nested PCR techniques. Until now HCV-PCR has been complicated, time-consuming and costly, and therefore not suitable for routine diagnostics. The PCR method described here is easy to perform , fast and cost-effective.