Serological Diagnosis of Autoimmune Bullous Skin Diseases

Abstract

Autoimmune bullous dermatoses (AIBD) encompass a variety of organ-specific autoimmune diseases that manifest with cutaneous and/or mucosal blisters and erosions. They are characterized by autoantibodies targeting structural proteins of the skin, which are responsible for the intercellular contact between epidermal keratinocytes and for adhesion of the basal keratinocytes to the dermis. The autoantibodies disrupt the adhesive functions, leading to splitting and blister formation. In pemphigus diseases, blisters form intraepidermally, whereas in all other disease types they occur subepidermally. Early identification of autoimmune bullous dermatoses is crucial for both treatment and prognosis, particularly as regards tumor-associated disease entities. The diagnosis is based on clinical symptoms, histopathology, direct immunofluorescence to detect antibody/complement deposits, and the determination of circulating autoantibodies. The identification of various target antigens has paved the way for the recent development of numerous specific autoantibody tests. In particular, optimized designer antigens and multiplex test formats for indirect immunofluorescence and ELISA have enhanced and refined the laboratory analysis, enabling highly efficient serodiagnosis and follow-up. This review elaborates on the current standards in the serological diagnostics for autoimmune bullous dermatoses.

Keywords: ELISA; autoantibody; autoimmune bullous dermatosis; biochip; indirect immunofluorescence; pemphigoid; pemphigus; serology.

Figure 1

Schematic presentation of human skin, depicting the targets of autoantibodies in autoimmune bullous dermatoses, reproduced from Gosink and Schlumberger, MEDLAB Magazine 2016 (1) with permission of MEDLAB Magazine. Top circle: antigenic structural components of desmosomes, which interconnect the cytoskeletons of neighboring keratinocytes in the epidermis. Bottom circle: antigenic structural components of hemidesmosomes, which anchor the cells of the epidermal stratum basale in the underlying basal lamina at the dermal-epidermal junction.

Figure 2

Indirect immunofluorescence staining of different tissue substrates in autoimmune bullous dermatoses. (A,B) Monkey esophagus, (C,D) monkey salt-split skin, (E) rat urinary bladder and (F) monkey liver. Graphics and annotations on the right indicate the staining patterns and possible underlying reactivities, reproduced and modified from Euroimmun customer leaflet, with permission of Euroimmun, Germany.

Figure 3

Detection and differentiation of autoantibodies in autoimmune bullous dermatoses using monospecific substrates for BIOCHIP-based indirect immunofluorescence, reproduced (in part) from Gosink and Schlumberger, MEDLAB Magazine 2016 (1) and from Gosink, MEDLAB Magazine 2013 (148) with permission of MEDLAB Magazine. (A–D) Substrates based on human embryonic kidney (HEK293) cells expressing recombinant immunodominant antigen domains: (A) Dsg1 (ectodomain), (B) Dsg3 (ectodomain), (C) BP230gC (globular C-terminal domain), (D) type VII collagen (NC1 domain). (E,F) Substrates generated by spotting purified recombinant protein: (E) BP180-NC16A-4X (tetrameric NC16A domain), (F) GAF-3X (trimeric deamidated gliadin-analogous fusion peptide).

Figure 4

BIOCHIP mosaics for simultaneous screening and monospecific confirmation of autoantibodies using indirect immunofluorescence, modified from Gosink, MEDLAB Magazine 2013 (148) with permission of MEDLAB Magazine. (A) “Dermatology Mosaic 7” (six substrates per reaction field). (B) “Dermatology Mosaic 11” (11 substrates per reaction field for extended analysis including paraneoplastic pemphigus and dermatitis herpetiformis). As indicated, the BIOCHIPs are coated with tissue sections (monkey esophagus, salt-split skin, liver, rat urinary bladder), HEK293 cells expressing recombinant antigens (Dsg1, Dsg3, BP230gC), or spots of purified recombinant antigen (BP180-NC16A-4X, GAF-3X). ^*HEp-2 and mock-transfected HEK293 cells serve as negative control substrates.