Monoclonal precipitating antibodies to porcine immunoglobulin M

Abstract

Fusion of splenic immunocytes from a porcine IgM-immunized BALB/c mouse with SP2/0 mouse myeloma cells resulted in 231 primary hybrids. Culture fluids of the primary hybrids were screened for antibody production by enzyme-linked immunosorbent assay (ELISA) against porcine IgM and by radial immunodiffusion versus porcine serum. Culture fluids of 10 of the primary hybrids were positive in IgM-ELISA and radial immunodiffusion. Six of these primary hybrids (1A11, 1D10, 2D7, 2E2, 3B11, and 5C9) were cloned, and ascitic fluids were produced using cloned primary hybrids. The monoclonal antibodies (Mabs) in ascitic fluids were characterized as to their reactivity with porcine immunoglobulin isotypes. All six Mabs had mouse IgG1, K isotype and were μ-chain specific as they formed single precipitin lines against porcine serum and porcine IgM and no lines against porcine IgG, IgA, and fetal porcine serum in immunodiffusion and immunoelectrophoresis. In indirect ELISA, all Mabs reacted with porcine serum, porcine IgM, and μ-chains but did not react with porcine IgG, IgA, or light chains. All six Mabs were species-specific and recognized either of two antigenic regions of μ-chain. These Mabs have been successfully used to detect IgM-containing cells in tissue sections, to detect IgM in serum, and to quantitate surface membrane IgM-bearing cells in peripheral blood.