Preparation and functional properties of polyclonal and monoclonal antibodies to murine MD-1.
Rabbits, rats and hamsters were immunized with KLH-coupled synthetic peptide sequences of the murine MD-1 molecule. Serum from immunized animals bound in Western gels to a 25 KDa protein extracted from LPS stimulated mouse spleen cells, as did a rat hybridoma (SH1.2.47) prepared from peptide-immunized rats. CHO cells transfected with a plasmid cDNA construct encoding murine MD-1, the target antigen for the antibodies in question, were also stained (in FACS) by the same antibodies. Patching and capping of the antigen(s) detected by any one of these sera abolished binding of all antibodies in subsequent FACS analysis, consistent with the hypothesis that they all detected the same antigen. In a final study to assess the possible involvement of MD-1 in regulation of cell activation for cytokine production following allostimulation, we found that all of the antibodies inhibited IL-2 and IFNgamma production, while enhancing IL-4 and IL-10 production, in mixed leukocyte reactions (MLR) in vitro.
Hadidi
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Transplant Research Division, CCRW 2-855 The Toronto Hospital, 200 Elizabeth Street, Ont., M5G 2C4, Toronto, Canada.
Yu
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Chen
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Gorczynski
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D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry / Q000378:metabolism*; D000954:Antigens, Surface / Q000276:immunology* / Q000378:metabolism; D001666:Binding Sites, Antibody; D015153:Blotting, Western; D016466:CHO Cells; D002460:Cell Line; D006224:Cricetinae; D016207:Cytokines / Q000096:biosynthesis; D005434:Flow Cytometry; D007106:Immune Sera / Q000737:chemistry / Q000378:metabolism*; D008070:Lipopolysaccharides / Q000494:pharmacology; D007959:Lymphocyte Culture Test, Mixed; D008212:Lymphocyte Depletion; D008562:Membrane Glycoproteins; D051379:Mice; D008809:Mice, Inbred C3H; D008810:Mice, Inbred C57BL; D008969:Molecular Sequence Data; D011817:Rabbits; D051381:Rats; D011917:Rats, Inbred Lew; D013154:Spleen / Q000166:cytology; D013194:Staining and Labeling; D013601:T-Lymphocytes / Q000276:immunology / Q000378:metabolism; D014162:Transfection
D000911:Antibodies, Monoclonal; D000954:Antigens, Surface; D016207:Cytokines; D007106:Immune Sera; D008070:Lipopolysaccharides; C113553:Ly86 protein, mouse; D008562:Membrane Glycoproteins