Detection of the 170-kDa bullous pemphigoid antigen by immunoprecipitation.
There has been controversy concerning the nature of the bullous pemphigoid (BP) antigen: immunoprecipitation identified BP antigen as a single, unique 230-kDa protein, whereas immunoblot analysis showed multiple antigen molecules, mainly 230- and 170-kDa proteins. In this study, to further characterize the 170-kDa protein, we have examined whether the 170-kDa protein is detected by immunoprecipitation. Extracts of human squamous cell carcinoma cells revealed the 170-kDa protein with immunoblot analysis. Although the conventional immunoprecipitation detected only the 230-kDa protein, some BP sera that detected the 170-kDa protein with immunoblotting also precipitated the 170-kDa protein with our modified immunoprecipitation, in which the cells were extracted with 1% sodium dodecylsulfate (SDS) buffer and reacted with the sera under reduced SDS concentration. The 170-kDa protein-specific BP sera clearly showed hemidesmosomal plaque staining with immunofluorescence of cultured cells. These results indicate that the 170-kDa protein is indeed one of the BP antigens and that the 230- and 170-kDa BP antigens are integrated in different ways in hemidesmosomes.
Ebihara
T
T
Department of Dermatology, Keio University School of Medicine, Tokyo, Japan.
Hashimoto
T
T
Kudoh
J
J
Gamou
S
S
Shimizu
N
N
Nishikawa
T
T
D001324:Autoantigens / Q000032:analysis*; D002352:Carrier Proteins; D003094:Collagen; D003598:Cytoskeletal Proteins; D000071821:Dystonin; D006801:Humans; D015151:Immunoblotting; D009419:Nerve Tissue Proteins; D024041:Non-Fibrillar Collagens; D010391:Pemphigoid, Bullous / Q000276:immunology*; D011233:Precipitin Tests / Q000379:methods; D012967:Sodium Dodecyl Sulfate; D014407:Tumor Cells, Cultured; D000097805:Collagen Type XVII
D001324:Autoantigens; D002352:Carrier Proteins; D003094:Collagen; D003598:Cytoskeletal Proteins; D000071821:Dystonin; D009419:Nerve Tissue Proteins; D024041:Non-Fibrillar Collagens; D012967:Sodium Dodecyl Sulfate; D000097805:Collagen Type XVII; C486268:DST protein, human