C. Milstein

Monoclonal Antibodies as Tools to Analyze the Serological and Genetic Complexities of Major Transplantation Antigens

The rat MHC resembles that of other species in displaying extensive polymorphism for a variety of MHC-characteristic functions: antigens detected by serological assays, antigens detected by cellular assays such as the MLR, GVH and CML, and immune response genes for a variety of cellular and non-cellular antigens (Gunther & Stark 1977, Gasser 1977). Nevertheless very little is known about the genetic structure of the region because of the shortage of laboratory recombinants. The present study grew out of the realisation that the high resolving power of monoclonal antibodies against complex polymorphic antigens could compensate for lack of resolution at the genetic level. Granted suitable monoclonal alloantibodies, the number and antigenic structure of the polymorphic molecules specified by the MHC can in principle be examined with greater precision than is possible by analysis of recombinants using planned immunizations and absorptions of conventional sera. This review describes the preparation of monoclonal antibodies by fusion of spleen cells from alloimmunized rats with mouse plasmacytoma cells and some results of an analysis of the properties of these antibodies. Some preliminary data have been published elsewhere (Galfre et al. 1977, Howard et al. 1978).

Monoclonal antibodies and cell surface antigens

Antibody chains are encoded in three gene clusters containing genes for the variable and constant regions. V and C genes are separated in germ line and during differentiation a rearrangement takes place. But even after this rearrangement the V and C coding sequences are not contiguous. A final splicing must take place in committed cells between the transcription of a discontinuous V-and C-region DNA and the expression of a continuous mRNA coding for an antibody chain. Analysis by cell fusion indicates that the splicing is cis. When two antibody-producing cell lines are fused, the resulting hybrids express the two antibodies that characterize the parental lines. Permanent cell lines producing antibody of predefined specificity have now been derived in this way. Spleen cells from hyperimmunized donors are fused with myeloma cells and a proportion of the hybrids that are established synthesize and secrete antibodies directed against the immunogen. The heterogeneous cell population can be cloned and propagated. This is a potent way of producing monospecific antibodies to complex antigens such as cell membranes and transplantation antigens. Monoclonal xenogeneic antibodies to rat cell-surface membranes have proved very valuable for characterizing and separating rat lymphocyte subpopulations. In more recent experiments, monoclonal xenogeneic antibodies to mouse and human cell-surface antigens have also been produced which permit the characterization of the hitherto undescribed differentiation antigens.