A Quantitative Assay for Cell-Bound Antibody Using a Radiolabeled Coombs-Type Antiserum to a γG Myeloma Protein

Abstract

Abstract Harder and McKhann have introduced an isotopic antiglobulin technique for measuring antibodies to cell-surface antigens (1). Sparks et al. have refined the technique to detect alloantibodies formed after first-set rejection of skin grafts (2). Further, Ting and Herberman have used it to study the humoral antibody response to a tumor-specific cell surface antigen (3). Two additional refinements are reported in this paper. The first is the preparation of a Coombs antiserum by immunizing rabbits with a highly purified mouse myeloma protein of γG type, thereby producing highly specific antibodies which cross-react with normal mouse γG globulin (4). The second is the purification of these antibodies by use of an insoluble antigen complex. The resulting purified Coombs antibody is radiolabeled and binds avidly to cell surfaces coated with mouse γG antibody. A very desirable end result, namely direct proportionality between the quantity of mouse antibody incubated with the cells and the amount of radiolabeled Coombs serum bound to the cell surface, has been obtained.