Characterization of monoclonal antibodies directed against swine leukocytes

Abstract

Hybridomas were produced from fusions of the SP 2 0 mouse myeloma with splenic cells from: 1) an outbred Sprague Dawley rat immunized with swine peripheral blood mononuclear (PBM) cells; 2) a ( CBA NDub × BALB c Dub ) F 1 mouse immunized with concanavalin A (Con A) activated swine PBM cells and 3)a ( BALB c Dub × C3H He Dub )F 1 mouse immunized with swine thymocytes. The resulting supernatants were screened by a microcytotoxicity assay for activity against swine PBM cells. Four hybridomas (MSA1, MSA2, MSA3 and MSA4) were selected, cloned and characterized by their cell reactivity and effect on mitogenic assays. MSA1 and MSA2 belong to the rat IgG2b subclass. MSA3 and MSA4 are of the mouse IgG2a subclass. These monoclonal antibodies reacted in the following manner: MSA1 with monocytes, granulocytes, red blood cells and bone marrow cells; MSA2 with subset of T cells; MSA3 with B cells and subsets of T cells and monocytes (class II molecule) and MSA4, a pan-T cell reagent (E-rosette receptor). The involvement of the various cell types reactive to the different monoclonal antibodies in the mitogenic response of swine PBM cells to Con A, phytohemagglutinin (PHA) or pokeweed mitogen (PWM) was investigated by cellular depletion with monoclonal antibody plus complement. Cellular depletion of PBM cells with the following monoclonal antibodies plus complement treatment resulted in: 1) MSA1, almost total reduction in the mitogenic response to low doses of Con A or PWM; 2) MSA2, partial reduction in the proliferative responses to any concentration of Con A, PHA or PWM; 3) MSA3, partial reduction in proliferative responses to low concentrations of Con A or PWM and 4) MSA4, total elimination of any proliferative response to Con A, PHA or PWM.