A NOVEL STRATEGY FOR GENERATING MONOCLONAL-ANTIBODIES FROM SINGLE, ISOLATED LYMPHOCYTES PRODUCING ANTIBODIES OF DEFINED SPECIFICITIES

We report a novel approach to the generation of monoclonal antibodies based on the molecular cloning and expression of immunoglobulin variab le region cDNAs generated from single rabbit or murine lymphocytes tha t were selected for the production of specific antibodies. Single cell s secreting antibodies for a specific peptide either from gp116 of the human cytomegalovirus or from gp120 of HIV-1 or for sheep red blood c ells were selected using antigen-specific hemolytic plaque assays. She ep red blood cells were coated with specific peptides in a procedure a pplicable to any antigen that can be biotinylated. Heavy- and light-ch ain variable region cDNAs were rescued from single cells by reverse tr anscription-PCR and expressed in the context of human immunoglobulin c onstant regions. These chimeric murine and rabbit monoclonal antibodie s replicated the target specificities of the original antibody-forming cells. The selected lymphocyte antibody method exploits the in vivo m echanisms that generate high-affinity antibodies. This method can use lymphocytes from peripheral blood, can exploit a variety of procedures that identify individual lymphocytes producing a particular antibody, acid is applicable to the generation of monoclonal antibodies from ma ny species, including humans.