ANTIGEN RETRIEVAL IN IMMUNOFLUORESCENT TESTING OF BULLOUS PEMPHIGOID

Direct immunofluorescence is an immunopathological technique frequentl y utilized for diagnosis of vesiculobullous disease such as bullous pe mphigoid. Fresh-frozen tissue is required for immunofluorescent testin g, making retrospective analysis difficult. In this study, we compared two methods of antigen retrieval in formalin-fixed, paraffin-embedded skin tissue from patients with bullous pemphigoid to determine if arc hival tissue, after use of an unmasking antigen, can be substituted fo r fresh-frozen tissue in the immunopathological study of skin. Paraffi n-embedded tissue blocks from patients with bullous pemphigoid and pat ients with eosinophilic spongiotic dermatitis as the prodromal stage o f bullous pemphigoid were obtained. Sections were mounted on poly-L-ly sine-coated slides and the slides were deparaffinized. The methods of antigen retrieval included incubation with trypsin (0.1%) and microwav e irradiation in urea (6 M). Antigen retrieval was followed by indirec t immunofluorescence. Microwave irradiation was more effective in anti gen retrieval than was incubation with trypsin (0.1%). Microwave irrad iation in urea (6 M) produced more intense immunofluorescent staining than did trypsinization. Overall, positive basement membrane zone immu nofluorescent staining was found in 60% of patients with a diagnosis o f classical bullous pemphigoid and in 50% of patients with eosinophili c spongiotic dermatitis as the prodromal stage of bullous pemphigoid. Although the frozen-tissue method appeared more effective than the ant igen-retrieval method in immunofluorescent testing of skin, the antige n-retrieval method can certainly be considered an option in retrospect ive studies. Antigen retrieval may be particularly advantageous in pat ients with eosinophilic spongiotic dermatitis in whom the diagnosis of bullous pemphigoid may not be suspected initially.