Biochemical analysis of ligand-induced receptor patching and capping using a novel immunolactoperoxidase iodination technique.
A novel approach for the analysis of membrane proteins involved in ligand-induced surface receptor patching and capping is described. The technique is based on the use of immunolactoperoxidase (immuno-LPO) conjugates which catalyze the iodination of those surface proteins with available tyrosine groups that are located in the immediate vicinity of the patch or cap of a particular antigen. We have used the patching and capping of the H-2 (histocompatibility) antigen on mouse thymocytes to illustrate this method. However, this technique should be generally applicable to any cell surface proteins which can be induced to form patches or caps by a specific ligand. Cytochemical analysis indicates that the immuno-LPO conjugates induce the same patching and capping of the H-2 antigen as does the unconjugated antibody. Biochemical analysis of the 125I-labeled proteins by SDS polyacrylamide gel electrophoresis indicates that a large membrane protein (mol wt of approximately 200,000 daltons) is closely associated with H-2 patches and caps. Since a number of other prominent membrane proteins are not labeled by this procedure, selective redistribution of certain surface proteins must be occurring during H-2 antibody-induced patching and capping.
D000818:Animals; D000906:Antibodies; D000936:Antigen-Antibody Complex; D004591:Electrophoresis, Polyacrylamide Gel; D006183:H-2 Antigens / Q000276:immunology*; D007124:Immunoenzyme Techniques; D007152:Immunologic Capping; D008565:Membrane Proteins / Q000032:analysis; D051379:Mice; D011948:Receptors, Antigen, T-Cell / Q000276:immunology*; D013601:T-Lymphocytes / Q000276:immunology*
D000906:Antibodies; D000936:Antigen-Antibody Complex; D006183:H-2 Antigens; D008565:Membrane Proteins; D011948:Receptors, Antigen, T-Cell