Production of murine monoclonal antibodies against a liver-specific, species cross-reactive antigen in the liver-specific lipoprotein (LSP) preparation

Abstract

Previous attempts in several laboratories have failed to produce murine monoclonal antibodies (MAbs) against liver-specific, species-cross-reactive, cell surface-expressed antigens in the normal liver preparation known as liver-specific membrane lipoprotein (LSP). In the present study, BALB/c mice were pretreated with a single dose of cyclophosphamide (20 mg/kg), hyperimmunized with human LSP and hybridomas produced by fusion of spleen cells from these mice with murine myeloma (P3-NS1-Ag4-1) cells. To bias selection in favour of MAbs reacting with species cross-reactive epitopes, hybridoma supernatants were screened by ELISA against rabbit LSP. From 70 stable hybridomas, four MAbs were obtained that react with rabbit LSP. One is an IgM antibody and the other three are of IgG2a class. All four react with the hepatic asialo-glycoprotein receptor (HL), a liver-specific, species-cross-reactive component that is normally expressed on the surfaces of hepatocytes. Using a rapid screening technique (an ‘additive’ ELISA), preliminary evidence was obtained indicating that these four MAbs between them recognise three different epitopes on the HL molecule. The results suggest that this is a viable approach for the production of MAbs against autoantigens to which autoreactivity may normally be suppressed.