Antibodies directed against monomorphic and evolutionary conserved self epitopes may be generated in 'knock-out' XXXX . Development of monoclonal antibodies directed against monomorphic MHC class I determinants.

@entity1 @entity8022 @entity19 @entity73224 @entity10062 @entity8200 @entity12812 @entity35 @entity291
@entity12812 ( @entity12812 ) gene 'knock-out' @entity19 ( @entity73224 ) were primed with purified H-2Kb and H-2Db molecules and spleen cells from immunized @entity19 were used to generate monoclonal antibody secreting B-cell hybridomas. Approximately 0.2% of the Ig-secreting primary microcultures contained H-2b binding antibodies. Three stable anti-MHC class I (MHC-I) antibody secreting hybridoma clones were established and subcloned. All three MoAbs precipitated radiolabelled @entity8200 molecules as analysed by SDS PAGE, and all three MoAbs stained H-2b, @entity8022 , as well as H-2k cells by @entity10062 analysis. The MoAbs stained to two @entity12812 loss mutant cell lines, C4.4-25- and R1E, suggesting that some MHC-I heavy chain is exported to the cell surface even in the absence of endogenous @entity12812 . Staining of @entity19 cell lines kept under serum-free culture conditions was strongly influenced by the addition of @entity291 or human serum as a source of exogenous @entity12812 suggesting that xenogeneic @entity12812 affects the conformation of class I molecules. Furthermore, all three MoAbs strongly stained the peptide transporter deficient cell line, RMA-S, when cultured at 26 degrees C, however, staining was reduced five-fold when RMA-S cells were cultured at 37 degrees C. In total, these observations suggest that the MoAbs recognize conformational, presumably @entity12812 and peptide dependent, self epitopes on MHC-class I. One of the three MoAbs stained @entity35 blood mononuclear blood cells (BMC), all three MoAbs stained hamster BMC, whereas two of the MoAbs stained @entity1 cells. These data suggest that the MoAbs recognize determinants which are conserved between species. All three antibodies strongly inhibited the development of CTLs generated in an allogeneic one-way MLC, provided that the MoAbs were present during the first 24 h of culture. It is concluded that MoAbs reacting with monomorphic self epitopes may be generated using animals deleted of the gene of interest. The implications may be far reaching since such MoAbs potentially identify evolutionary conserved and physiologically important epitopes.
