Bullous pemphigoid sera react specifically with various domains of BP230, most frequently with C-terminal domain, by immunoblot analyses using bacterial recombinant proteins covering the entire molecule.
By immunoblot analyses of normal human epidermal extracts, the 230 kDa bullous pemphigoid antigen (BP230) is recognized by most bullous pemphigoid sera. By polymerase chain reaction using keratinocyte cDNA library as a template, we successfully amplified 3 cDNAs of about 3 kb, which covered whole human BP230 molecule. By inserting the cDNAs into bacterial expression vector pGEX, we prepared 3 different recombinant glutathione-S-transferase-fusion proteins, which roughly presented N-terminal domain, central rod domain and C-terminal domain of BP230. By immunoblotting using these 3 recombinant proteins, we demonstrated that the majority of bullous pemphigoid sera reacted clearly with multiple recombinant proteins of BP230, most frequently with C-terminal domain. We also examined sera of pemphigus vulgaris, pemphigus foliaceus and herpetiform pemphigus that showed BP230-like protein band by immunoblotting of epidermal extracts, as well as paraneoplastic pemphigus, for reactivity with the 3 recombinant proteins. In the study, we found that only very few of these non-bullous pemphigoid sera reacted with some of the recombinant proteins. These results indicate that the BP230 is specifically reacted by bullous pemphigoid sera, and that the immunoblotting using the BP230 recombinant proteins should be a useful tool for the diagnosis of bullous pemphigoid.
Hamada
T
T
Department of Dermatology, Kurume University School of Medicine, 67 Asahi-machi, Kurume, Fukuoka 830-0011, Japan.
Nagata
Y
Y
Tomita
M
M
Salmhofer
W
W
Hashimoto
T
T
D000911:Antibodies, Monoclonal / Q000276:immunology; D001324:Autoantigens / Q000737:chemistry* / Q000276:immunology*; D001327:Autoimmune Diseases / Q000097:blood / Q000276:immunology; D001426:Bacterial Proteins; D001769:Blood / Q000276:immunology; D002352:Carrier Proteins; D003094:Collagen / Q000737:chemistry* / Q000276:immunology*; D003598:Cytoskeletal Proteins; D000071821:Dystonin; D006801:Humans; D015151:Immunoblotting; D009419:Nerve Tissue Proteins; D024041:Non-Fibrillar Collagens; D010391:Pemphigoid, Bullous / Q000097:blood* / Q000276:immunology; D011993:Recombinant Fusion Proteins; D012867:Skin / Q000276:immunology; D012872:Skin Diseases, Vesiculobullous / Q000097:blood / Q000276:immunology; D000097805:Collagen Type XVII
D000911:Antibodies, Monoclonal; D001324:Autoantigens; D001426:Bacterial Proteins; D002352:Carrier Proteins; D003094:Collagen; D003598:Cytoskeletal Proteins; D000071821:Dystonin; D009419:Nerve Tissue Proteins; D024041:Non-Fibrillar Collagens; D011993:Recombinant Fusion Proteins; D000097805:Collagen Type XVII; C486268:DST protein, human