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Title: Monoclonal antibodies to Ia glycoproteins from rat thymus and spleen
Author: McMaster, William Robert
ISNI:       0000 0001 3625 8298
Awarding Body: University of Oxford
Current Institution: University of Oxford
Date of Award: 1979
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A fraction of rat thymocyte la glycoproteins was prepared by lentil lectin affinity chromatography and gel filtration in deoxycholate. Mice were immunized with these glycoproteins and cell lines secreting monoclonal antibody produced by cell fusion. Antibody from four cell lines, MRC OX 3, 4, 5 and 6, reacted with the thymus la glycoproteins. Analysis using a fluorescence activated cell sorter showed that these monoclonal antibodies bound to the majority of peripheral B lymphocytes, a subpopulation of thymocytes (18%) but did not significantly bind to peripheral T lymphocytes. MRC OX 3 antibody detected a polymorphic antigenic determinant found on rat strains of the RT1 haplotype \ and y while MRC OX 4, 5 and 6 antibodies detected an antigenic determinant common to the rat strains of the RT1 haplotype a, c, 1, n, y. All four antibodies bound to mouse spleen cells and all detected polymorphic determinants. Studies on recombinant mouse strains suggested that the determinants recognized were coded by the I-A subregion of the tt-2 complex. MRC OX 3 antibody also cross reacted with a polymorphic determinant on human lymphoblastoid cell lines. This antibody bound to cells which expressed the HLA-DRw specificities DRwl, DRw2 or DRw6 but did not bind to those cells which only expressed other HLA-DRw specificities. No significant binding of MRC OX 3 or 4 antibody to either normal or mitogen stimulated human peripheral leukocytes c:uld be detected. MRC OX 4 antibody was used for affinity chromatography to purify la glycoproteins from rat spleen. The rat spleen la antigen was composed of two non-covalently bonded polypeptides of apparent molecular weight (unreduced) 30,000 and 24,000 as determined by polyerjryl amide gel electrophoresis in sodium dodecyl sulfate. The spleen la antigen had molecular characteristics indistinguishable from the thymus la glycoproteins. The purified spleen la antigen partially inhibited the binding to lymph nod« cells of rat alloantibodies which detect la antigens linked to the RTi complex. The la absorbed alloantibodies still detected antigens found only on peripheral B lymphocytes. This binding could be completely inhibited by a further absorption with lymph node glycoproteins depleted of the MRC OX 4 la antigen. These results suggested that there were at least two separate rat la antigens. One antigen was detected by MRC OX 4 antibody while the remaining antigens were detected by the MRC OX 4 la absorbed alloantibodies. From the same immunization another cell line was prepared called MRC OX 2 which secreted monoclonal antibody to a previously undefined thymus glycoprotein of 60,000 molecular weight. The MRC OX 2 antigen was found on all thymocytes and on brain tissue in equal amounts on the basis of wet tissue weight, and was also found on peripheral B lymphocytes in smaller amounts. Spleen, kidney and liver tissue had less than 10% of the antigen as compared to thymocyte or brain tissue.
Supervisor: Williams, Anthony Ffoulkes Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available
Keywords: Monoclonal antibodies ; Glycoproteins