Pemphigus is a human disease that causes extensive blistering of the skin. This blistering is related to a loss of epidermal cell cohesion and is accompanied by circulating autoantibodies that stain epidermal cell surfaces, as shown by immunofluorescence microscopy. One of the major components involved in epidermal cell cohesion is the desmosome. The pathological changes that accompany pemphigus led us to determine whether the autoantibodies are specific for desmosomes. Incubation of cultured mouse keratinocytes in medium containing pemphigus antiserum leads to cell separation at cell-cell contact sites, which possess desmosomes. Tissue sections of mouse skin processed for indirect immunofluorescence, using pemphigus antiserum or a rabbit antiserum directed against components of desmosomes, show similar punctate cell-surface staining patterns within the epidermis. Cultured mouse keratinocytes possessing well-defined intermediate filament bundles (tonofilaments) and desmosomes were processed for double indirect immunofluorescence, using a monoclonal antibody directed against mouse skin keratin and either pemphigus antiserum or the desmosome antiserum. The keratinocytes exhibit a complex system of keratin-containing tonofilaments. Tonofilaments in contacting cells are separated by thin dark bands at the cell surface, which correspond precisely to desmosomal plaques seen by phase-contrast microscopy. These bands specifically stain with both pemphigus antiserum and the desmosome antiserum. Double indirect immunofluorescence of the cultured mouse keratinocytes, using pemphigus antiserum and the desmosome antiserum, reveals that the pemphigus autoantibodies stain the same areas of cell-cell contact as the desmosome antibodies. Our evidence supports the idea that pemphigus blisters form, at least in part, from a specific antibody-induced disruption of desmosomes in the epidermis.