There are many known methods of dermo-epidermal separation for the investigation of autoimmune blistering diseases. Investigators should select a proper method since many differences exist preservation of antigenicity. In order to determine the stabilization of antigenirity by different separation methods, we have separated dermo-epidermal junction by means of 1M s;ilt, 56C PBS, 20mM EDTA and dispase. Indirect immunofluarescence and immunoblotting were performed on each specimen with sera of patients with pemphigus vulgaris, pemphigus foliaceus, paraneoplastic pemphigus, bullous pemphigoid and epidermolysis bullosa acquisita. The results are as follows : 1. In indirect immunofluorescence study of pemphigus group, best, result were obtained when normal skin without dermo-epidermal separation was used. Dispase well preserved antigenicity of pemphigus after dermo-epidermal separation, but no differences were noted in antigenicity stabilization among separation mehods by immunoblotting. 2. In indirect immunofluorecence study for differentiation of bullous pemphigoid and epidermolysis bullosa acquisita, we recommend EDTA and dispase methods in addition to 1M salt induced skin separation that have been most popularly used. 3. Results of the immunoblotting of bullous pemphigoid showed that 1M salt, EDTA and heat preserved the antigenicity well but the antigenicity was lost by dispase. 4. Results of the immunoblotting of epidermolysis bullosa acquisita she wed that antigen did not exist in epidermal extract. 5. Antigen preservation according to the different methods of demo-epidermal separation was not identical between indirect immunofluorescence and immunoblotting (Kor J Dermatol 1993; 31 (1): 19-27)