Cell membrane antigen isolation with the staphylococcal protein A-antibody adsorbent.
Procedures are detailed for the rapid isolation of representative cell membrane antigens with protein A-bearing staphylococci as an adsorbent for IgG antibodies complexed with the antigens. Cell surface membrane proteins were radioiodinated and solubilized in nonionic detergent. Specific antisera were subsequently added and the immune complexes precipitated by addition of the staphylococcal adsorbent and low speed centrifugation. The antigens isolated included surface immunoglobulins from mouse and human lymphocytes, human beta-microglobulin and HL-A alloantigens, mouse H-2 alloantigens, and the murine leukemia virus glycoprotein gp 70. Rabbit, sheep, goat, and mouse antisera were all effective for the specific phase of the precipitation reaction. The surface membrane immunoglobulins of mouse splenic lymphocytes and human peripheral blood lymphocytes differed with respect to class composition and protein A reactivity. Mouse lymphocyte surface immunoglobulins were nonreactive with protein A, whereas a high proportion of human lymphocyte surface immunoglobulins of different classes bound directly to the staphylococci. In sequential immunoprecipitation studies the prior isolation of one antigen had no appreciable effect on the subsequent recovery of another antigen. Adsorption of antigen-antibody complexes is quantitative when protein A sites are provided in excess over antiserum IgG sites, and this obviates the need for equivalence point titrations for optimal precipitation necessary with alternative double antibody techniques.
D000818:Animals; D000936:Antigen-Antibody Complex; D000941:Antigens / Q000302:isolation & purification*; D001426:Bacterial Proteins / Q000276:immunology*; D002462:Cell Membrane / Q000276:immunology*; D006041:Goats; D006680:HLA Antigens / Q000302:isolation & purification; D006649:Histocompatibility Antigens / Q000302:isolation & purification; D006801:Humans; D007122:Immunoelectrophoresis; D007163:Immunosorbent Techniques; D007164:Immunosorbents / Q000276:immunology; D009052:Leukemia Virus, Murine / Q000276:immunology; D051379:Mice; D011817:Rabbits; D011947:Receptors, Antigen, B-Cell / Q000302:isolation & purification; D012756:Sheep; D013154:Spleen / Q000276:immunology; D013210:Staphylococcus; D014764:Viral Proteins / Q000302:isolation & purification
D000936:Antigen-Antibody Complex; D000941:Antigens; D001426:Bacterial Proteins; D006680:HLA Antigens; D006649:Histocompatibility Antigens; D007164:Immunosorbents; D011947:Receptors, Antigen, B-Cell; D014764:Viral Proteins