CHARACTERIZATION OF A PANEL OF MONOCLONAL-ANTIBODIES RAISED AGAINST RECOMBINANT HCV CORE PROTEIN

Hepatitis C virus (HCV) has as yet no practical culture system so any antigen or antibody studies must be carried out using recombinant anti gens. In this study, HCV core sequence was amplified by PCR, inserted into pRSET, and expressed in E. coli. The resultant core protein was p urified using nickel affinity chromatography which bound the six histi dine tag attached to the N-terminus of the protein. After elution in i midazole buffer, the core protein was used to immunise Balb/c mice and monoclonal antibodies against HCV core were raised. Six monoclonals w ere examined in a variety of assays. All of them recognised the p27 kD a protein which they were raised against and 2D2 and 3D7 recognised th e core component of an HCV Recombinant immunoblot Assay (RIBA). None o f the antibodies recognised the linear peptides in an Innolia HCV assa y. 2D2 showed cytoplasmic granular staining in 1-5% of cells in frozen sections of HCV infected livers. Cross-competition assays between the mselves and human anti-HCV core positive sera divided the antibodies i nto two main groups (I and II), with a sub-division of group into a an d b. Group I antibodies were unable to be inhibited by human anti-HCV sera whereas group II antibodies were inhibited by these sera (up to 6 2%), Epitopes recognised by all the monoclonals were probably conforma tional with the group I epitope being located within the first 105 ami no acids of the core sequence and the group II epitope between amino a cids 105 and 160. (C) 1996 Wiley-Liss, Inc.