Testing for cell surface forms of class II major histocompatibility complex antigens and Ii by radioiodination, biotinylation, and membrane immunofluorescence.
Antibodies to either Ii or class II major histocompatibility complex (MHC) antigens did not recognize cell surface forms of Ii in immunoprecipitates of cells that had been radioiodinated by the lactoperoxidase method, whereas they bound [35S]methionine metabolically labeled molecules. N-hydroxysuccinimidobiotin (NHS-B) and biotin hydrazide (B-H) were used to react more generally with cell surface proteins via amino groups and nitrene coupling, respectively. Each of these latter compounds labeled alpha and beta chains of class II MHC antigens as seen in Western-blotted, electrophoresed immunoprecipitates probed with 125I-labeled streptavidin but not Ii or its associated forms. Although tyrosine residues might have been inaccessible to radioiodination in carbohydrate-derivatized forms of Ii, the lack of Ii biotinylation in these controlled, sensitive studies was consistent with the view that Ii forms were not surface expressed, with the possible exception of the chondroitin sulfate-derivatized forms of Ii (Ii-CS).
Elliott
W L
WL
Department of Pharmacology, University of Massachusetts Medical School, Worcester 01655.
Sorli
C H
CH
Reisert
P S
PS
Humphreys
R E
RE
D000954:Antigens, Surface / Q000032:analysis*; D001710:Biotin / Q000031:analogs & derivatives; D002087:Butyrates / Q000494:pharmacology; D020148:Butyric Acid; D002460:Cell Line; D005455:Fluorescent Antibody Technique; D006680:HLA Antigens / Q000032:analysis* / Q000145:classification; D006801:Humans; D007457:Iodine Radioisotopes
D000954:Antigens, Surface; D002087:Butyrates; D006680:HLA Antigens; D007457:Iodine Radioisotopes; D020148:Butyric Acid; C053311:biotin hydrazide; D001710:Biotin