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In situ-bound antibodies eluted from the skin of patients with bullous pemphigoid are preferentially directed against the 230-kD bullous pemphigoid antigen.

Two bullous pemphigoid antigens (BPAs) have been identified: a 230-kD intracellular hemidesmosome-associated molecule and a 180-kD transmembrane hemidesmosome-associated molecule. Although patients with bullous pemphigoid (BP) have been shown to have circulating antibodies directed against one or both BPAs, the antigenic specificity of tissue-bound BP autoantibodies has not been studied. Because these skin-bound antibodies may play an important role in disease initiation, we sought to determine their antigenic specificity. In situ-bound BP antibodies were eluted with glycine from salt-split perilesional skin biopsy specimens and subjected to immunoprecipitation. Ten of 13 patients had glycine-eluted antibodies that recognized the 230-kD BPA, whereas four of 13 patients had glycine-eluted antibodies that recognized the 180-kD BPA. When these glycine-eluted BP skin specimens were sequentially eluted with urea, we found either antibodies of the same specificity or loss of reactivity to one of the BPAs, but no reactivity to any BPAs previously undetected with glycine elution. Sequential elution with sodium dodecylsulfate revealed no detectable BP antibodies. The heavy- and light-chain isotypes of the circulating, tissue-bound, and eluted BP antibodies were very similar, suggesting that we are not eluting certain subsets of BP antibodies. Sera from these patients contained circulating antibodies that recognized either one or both BPAs. Our observations demonstrate that in situ-bound antibodies eluted from the skin of patients with BP are preferentially directed against the 230-kD BPA.