AA3 is a novel antibody raised against human amnion, which reacts with the basement membrane of various epithelia of ectodermal origin. We used AA3 to examine the epidermal basement membrane zone in normal skin and different genetically determined types of epidermolysis bullosa (EB), by indirect immunofluorescence. AA3 staining was normal in dystrophic and simplex EB, but was markedly reduced in lesional and non-blistered skin in severe forms of junctional EB. In non-lethal junctional EB, the intensity of staining was variable and appeared to be inversely associated with disease severity, but did not correlate with demonstrable abnormalities of hemidesmosomes. AA3 binding was not reduced in pemphigoid lesions or normal suction blisters. It appeared to localize to the lamina lucida, but with different characteristics compared with antibodies to laminin and bullous pemphigoid antigen. These finding suggest that AA3 recognizes an antigen (or antigens) which may be involved in a primary biochemical defect in junctional EB. Moreover, this antibody may act as a new probe for this potentially lethal mechano-bullous disease.