Double immunofluorescence microscopy: a method for localizing immune deposits in skin diseases associated with linear...

Direct immunofluorescence microscopy has shown that a linear pattern of immunoglobulin and/or complement deposition at the cutaneous basement membrane zone is a characteristic feature in a number of acquired bullous diseases and is occasionally observed in systemic lupus erythematosus. Immunoelectron microscopy has shown the linear pattern of immunofluorescence may be produced by immune deposits located either above the basal lamina (in the lamina lucida) or below the basal lamina (in the upper dermis). Distinguishing between these sites of immune reactant deposition may be of value in differential diagnosis. In this study we report a double immunofluorescent method by which skin biopsies with linear IgG immunofluorescence due to deposits above the basal lamina (bullous pemphigoid) could be distinguished from biopsies with deposits beneath the basal lamina (bullous systemic lupus erythematosus and epidermolysis bullosa acquisita). When skin sections were treated sequentially with rhodamine-labeled anti-human IgG followed by fluorescein-labeled antilamina lucida (pemphigoid) antibody and examined by fluorescence microscopy, the following results were obtained. In biopsies with IgG deposits in the lamina lucida, a single green fluorescent band was observed. In tissues with subbasal lamina deposits, either parallel and contiguous bands of green and yellow-orange fluorescence or a single band of yellow-orange fluorescence was observed. The method is simpler, quicker, and less expensive than immunoelectron microscopy and should be a useful technique for evaluating skin diseases with linear immunofluorescence at the basement membrane zone.