Comparative study of autoantigens for various bullous skin diseases by immunoblotting using different dermo-epidermal separation techniques.

We investigated the reactivity of pemphigus vulgaris (PV), pemphigus vegetans, pemphigus foliaceus (Pf), Brazilian Pf, bullous pemphigoid (BP), and epidermolysis bullosa acquisita (EBA) sera with an immunoblot analysis using human epidermal and dermal extracts as a source of antigen. To obtain epidermal and dermal extracts three different dermo-epidermal separation methods were used: namely, ethylenediaminetetraacetic acid (EDTA) separation, heat separation, and dispase separation. All the 15 PV and the seven pemphigus vegetans sera demonstrated a 130-kDa PV antigen in epidermal extracts obtained by all the three methods. Furthermore, three PV sera also showed a 160-kDa Pf antigen, desmoglein. Ten of 14 Pf sera and six of 15 Brazilian Pf sera reacted with desmoglein in the same pattern in all the three epidermal extracts. Fifteen of the 22 BP sera showed reactivity with 230-kDa BP antigen in the same pattern in all the three epidermal extracts, whereas 14 BP sera detected the 180-kDa BP antigen in extracts of EDTA- and heat-separated epidermis but not in dispase-separated epidermal extract. Dermal extracts were obtained by EDTA- and heat-separated dermis, and all six EBA sera labelled a 290-kDa EBA antigen in both samples. These results suggest that heat-separated skin is as useful as EDTA-separated skin for detecting various autoantigens, but heat separation is preferable because the preparation time is shorter.