Skin explants from two lesional areas and four normal-appearing areas of four patients with benign familial chronic pemphigus (BFCP) were organ cultured with and without various reagents. After 24-h culturing of involved skin with medium only, dissociation of keratinocytes, which was also observed prior to culturing, was exacerbated, and the epidermis became edematous, with a large section detaching from the dermis. These phenomena were not suppressed even when betamethasone, retinol acetate, or camostat mesilate (serine protease inhibitor) was added to the medium. On the other hand, in the cultures of uninvolved skin explants with medium only, widened intercellular spaces were observed 24-48 h after initiation of culture, and dissociation of keratinocytes and acantholytic clefts became apparent after 72 h. Such culture-induced acantholysis was almost completely suppressed by the addition of betamethasone, but not suppressed by the addition of retinol acetate, EDTA, N-ethylmaleimaide, or pepstatin A. Camostat and SBTI incompletely suppressed the acantholysis. These findings suggest the possibility that steroid may reduce blistering and that an organ culture of non-lesional benign familial chronic pemphigus (BFCP) skin may be useful for clarifying the pathogenesis, as well as for discovering new drugs for the treatment of BFCP. Further experiments are required to clarify the role of serine proteases in the acantholysis in this disease.