title: Transgenic mice secreting coronavirus neutralizing antibodies into the milk.
abstract: Ten lines of transgenic mice secreting transmissible gastroenteritis coronavirus (TGEV) neutralizing recombinant monoclonal antibodies (rMAbs) into the milk were generated. The rMAb light- and heavy-chain genes were assembled by fusing the genes encoding the variable modules of the murine MAb 6A.C3, which binds an interspecies conserved coronavirus epitope essential for virus infectivity, and a constant module from a porcine myeloma with the immunoglobulin A (IgA) isotype. The chimeric antibody led to dimer formation in the presence of J chain. The neutralization specific activity of the recombinant antibody produced in transiently or stably transformed cells was 50-fold higher than that of a monomeric rMAb with the IgG1 isotype and an identical binding site. This rMAb had titers of up to 10(4) by radioimmunoassay (RIA) and neutralized virus infectivity up to 10(4)-fold. Of 23 transgenic mice, 17 integrated both light and heavy chains, and at least 10 of them transmitted both genes to the progeny, leading to 100% of animals secreting functional TGEV neutralizing antibody during lactation. Selected mice produced milk with TGEV-specific antibody titers higher than 10(6) as determined by RIA, neutralized virus infectivity by 10(6)-fold, and produced up to 6 mg of antibody per ml. Antibody expression levels were transgene copy number independent and integration site dependent. Comicroinjection of the genomic beta-lactoglobulin gene with rMAb light- and heavy-chain genes led to the generation of transgenic mice carrying the three transgenes. The highest antibody titers were produced by transgenic mice that had integrated the antibody and beta-lactoglobulin genes, although the number of transgenic animals generated does not allow a definitive conclusion on the enhancing effect of beta-lactoglobulin cointegration. This approach may lead to the generation of transgenic animals providing lactogenic immunity to their progeny against enteric pathogens.
authors.affiliation: Centro Nacional de Biotecnología, Consejo Superior de Investigaciones Científicas, Department of Molecular and Cell Biology, Campus Universidad Autónoma, Cantoblanco, Madrid, Spain.
authors.forename: I
authors.initials: I
authors.lastname: Sola
authors.forename: J
authors.initials: J
authors.lastname: Castilla
authors.forename: B
authors.initials: B
authors.lastname: Pintado
authors.forename: J M
authors.initials: JM
authors.lastname: Sánchez-Morgado
authors.forename: C B
authors.initials: CB
authors.lastname: Whitelaw
authors.forename: A J
authors.initials: AJ
authors.lastname: Clark
authors.forename: L
authors.initials: L
authors.lastname: Enjuanes
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis; D000914:Antibodies, Viral / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology; D001483:Base Sequence; D019556:COS Cells; D002460:Cell Line; D002461:Cell Line, Transformed; D018076:DNA, Complementary; D006801:Humans; D007070:Immunoglobulin A / Q000096:biosynthesis* / Q000235:genetics; D007132:Immunoglobulin Isotypes; D007137:Immunoglobulin alpha-Chains / Q000096:biosynthesis / Q000235:genetics; D007145:Immunoglobulin kappa-Chains / Q000096:biosynthesis / Q000235:genetics; D051379:Mice; D008822:Mice, Transgenic; D008892:Milk / Q000276:immunology*; D008969:Molecular Sequence Data; D009500:Neutralization Tests; D011994:Recombinant Proteins / Q000096:biosynthesis; D013552:Swine; D005760:Transmissible gastroenteritis virus / Q000276:immunology*; D014407:Tumor Cells, Cultured
chemical_list: D000911:Antibodies, Monoclonal; D000914:Antibodies, Viral; D018076:DNA, Complementary; D007070:Immunoglobulin A; D007132:Immunoglobulin Isotypes; D007137:Immunoglobulin alpha-Chains; D007145:Immunoglobulin kappa-Chains; D011994:Recombinant Proteins