TYPING OF HEPATITIS-C VIRUS ISOLATES BY DNA ENZYME-IMMUNOASSAY

Recently, at least six types of hepatitis C viruses (HCV) have been id entified. Different types of HCV appear to possess different pathogeni c properties and a different sensitivity to interferon treatment. Typi ng of HCV isolates may therefore be an important diagnostic procedure. We report on a new method for identification of HCV types 1a, 1b, 2a, 2b and 3a which are most prevalent in Europe, North America and Japan , The assay is based on a combination of two well established techniqu es, the polymerase chain reaction (PCR) and DNA enzyme immunoassay (DE IA). In the first step of the method a cDNA of about 250 bp correspond ing to the HCV core-region is amplified by nested PCR. The target cDNA is then hybridized to type-specific oligonucleotides fixed to a solid phase through an avidin-biotin bridge. The formed hybrids are detecte d by a standard ELISA using monoclonal antibodies reacting with double -stranded DNA, Typically, signal-to-noise (S/N) ratios between 18.2 an d 48.6 could be observed when different HCV types/subtypes were analyz ed by this method. The test was evaluated using cloned HCV cDNAs of kn own types and by sequence determination of some of the typed cDNAs. Ty ping of 115 isolates from Germany, Russia and Turkey revealed that sub type 1b (59-100%) and 1a (24-32%) are most prevalent in these countrie s.