Indirect immunofluorescence microscopy

The purpose of indirect immunofluorescence microscopy is to detect circulating antibodies in patient’s serum. For this purpose, an adequate substrate is necessary to visualize these antibodies. Monkey esophagus is the most widely used substrate for detecting circulating autoantibodies in patients with autoimmune bullous diseases. In all variants of pemphigus, antibodies show an epithelial cell surface pattern, resulting from present autoantibodies against the desmosomal molecules desmoglein 1 and/or 3. This pattern is also called chicken wire or honeycomb pattern. In pemphigoid, a linear deposition along the epithelial basement membrane can be observed, caused by autoantibodies against hemidesmosomes or their connecting proteins underneath.Human salt-split skin is a valuable substrate in the diagnosis of subepidermal autoimmune bullous diseases. Important antigens in the roof of salt-split skin are type XVII collagen (BP180) and BP230, whereas laminin 332, p200, and type IV collagen are situated in the floor of the blister. This implies that bullous pemphigoid, mucous membrane pemphigoid, pemphigoid gestationis, and lichen planus pemphigoides show staining of IgG on the epidermal side of the blister. On the other hand, anti-laminin 332 pemphigoid, anti-p200 pemphigoid, epidermolysis bullosa acquisita, and bullous SLE show staining on the dermal side.Other less used, but valuable substrates in some instances, are rat bladder and knock-out skin