Antibody to immunoselected L-cell antigens mimics stimulating activity of antibody to whole L cells.
A heterogeneous IgG antibody raised in rabbits in response to injections of whole L cells was used to identify and select relevant antigens in a nonionic detergent extract of L cells prelabeled with [35S]methionine by means of immunoprecipitation and immunoaffinity chromatography. When analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, the immunoprecipitate and immunoeluate contained far fewer protein bands than the whole cell extract but selectively retained a 42,000-MW protein species. In response to injections of the immunoprecipitate, rabbits produced a new antiserum which reacted predominantly with the 42,000-MW protein when reacted with L-cell proteins separated by sodium dodecyl sulfate-gel electrophoresis and transferred to nitrocellulose paper by the Western blot technique. The new antiserum (raised to the immunoprecipitate) and the original antiserum (raised to whole cells) were equipotent in stimulating calcium transport, phospholipid metabolism, and DNA synthesis in L cells. Binding of the IgG fractions of the two antisera displayed identical high affinity binding to L-cell surface antigens, with the same average association constant of 1.5 X 10(6) M-1. These studies have shown that an antiserum raised to whole L cells has a much narrower reactive spectrum with L-cell membrane antigens than might be imagined and has identified a 42,000-MW membrane protein as an important immunogen which itself elicits a potent immune response resulting in an antibody capable of mimicking the cell stimulatory properties of the original antiserum.
D000818:Animals; D000918:Antibody Specificity; D000937:Antigen-Antibody Reactions; D000954:Antigens, Surface / Q000276:immunology*; D004261:DNA Replication; D007120:Immunochemistry; D007700:Kinetics; D007739:L Cells / Q000276:immunology* / Q000378:metabolism; D008565:Membrane Proteins / Q000276:immunology*; D051379:Mice; D008970:Molecular Weight
D000954:Antigens, Surface; D008565:Membrane Proteins