A novel method for radiolabeling antigen-binding receptors of lymphocytes.
Antigen-binding receptor (ABR) molecules have been selectively radiolabeled and isolated from immunized chicken spleen cells. The specific radiolabeling of the receptors has been accomplished by utilizing a novel technique employing lactoperoxidase (LPO) covalently linked to antigen (Ag) for which human gammaglobulin was used. The cell surface ABRs were first bound to the Ag-LPO conjugates through specific recognition sites on the Ag portion of the conjugates. The bound LPO portions were then allowed to catalyze the radioiodination of the ABRs. After radiolabeling, cells were solubilized with detergents. ABRs still bound to Ag-LPO conjugates were directly isolated from the lysates via immunoaffinity chromatography utilizing an immunoaffinity reagent directed toward the antigen portion of the ABR-Ag-LPO complex. The radioactive materials isolated in this way were then analyzed via SDS-PAGE under reducing conditions. It appears that as expected, most of the specifically-labeled and isolated materials were immunoglobulin (Ig). Both the membrane-bound form of the heavy chain (mol. wt 77 K) as well as the secreted form (mol. wt 67 K) were detected, along with the light chain (mol. wt 25 K). An additional polypeptide of mol. wt 55 K was also selectively labeled and isolated along with the Ig. This may be a molecule closely associated with the membrane immunoglobulin on B-cell surface.
D000818:Animals; D001402:B-Lymphocytes / Q000276:immunology; D002645:Chickens; D002846:Chromatography, Affinity; D004591:Electrophoresis, Polyacrylamide Gel; D007457:Iodine Radioisotopes; D007553:Isotope Labeling / Q000379:methods; D007784:Lactoperoxidase; D008970:Molecular Weight; D011947:Receptors, Antigen, B-Cell / Q000032:analysis* / Q000276:immunology; D011961:Receptors, Fc / Q000276:immunology; D013154:Spleen / Q000276:immunology
D007457:Iodine Radioisotopes; D011947:Receptors, Antigen, B-Cell; D011961:Receptors, Fc; D007784:Lactoperoxidase