Epitope mapping of BP230 leading to a novel enzyme‐linked immunosorbent assay for autoantibodies in bullous pemphigoid

Abstract

Recombinant BP230-C3 is a suitable target antigen for the detection of serum autoantibodies against BP230.

Anti-Skin Specific Autoantibodies Detected by a New Immunofluorescence Multiplex Biochip Method in Patients with Autoimmune Bullous Diseases

“…Although recombinant proteins used in the multiplex biochip have been studied by other researches with ELISA methods [20,21,22,24,25,26,27,28,29,30], this is the first study on their use with an IIF-based biochip method.…”

“…In detail, each field of the slide is a mosaic of 6 substrates: (1) frozen primate esophagus section; (2) EU90 recombinant cells transfected with Dsg1 protein ectodomain (aa 1–569) [10]; (3) EU90 recombinant cells transfected with Dsg3 protein ectodomain (aa 2–640) [10]; (4) EU90 recombinant cells transfected with full-length BP230 protein (aa 1–2649) [24]; (5) EU90 recombinant cells transfected with C-terminal globular domain of the BP230 protein (aa 1875–2649) [24]; (6) microdrops of NC16A-4X BP180 free antigen (tetramer of the immunogenic NC16A noncollagenosus domain of the BP180 protein, aa 490–562, expressed in Escherichia coli ) [18]. …”

“…The four Euroimmun ELISA microplates were coated with: (1) the extracellular domain of Dsg1 (aa 1–548) [10] and (2) Dsg3 (aa 2–616) [10] – both proteins based on human cDNA were produced recombinantly in mammalian cells (HEK293); (3) biochemically purified C-terminal sequence (aa 2326–2649) [24] of the human 230-kDa BP antigen expressed in E. coli; (4) recombinant BP180-NC16A-4X, a tetramer (four copies fused to a carboxyterminal polyhistidine tag) of the immunogenic NC16A domain (aa 490–562) [18] of BP180, based on human cDNA, expressed in E. coli. …”

IgE-mediated mechanisms in bullous pemphigoid and other autoimmune bullous diseases

“…[14] BP230 is targeted in about 50% of patients, with the most immunogenic epitopes located in the globular C-terminal portion. [15,16] It has been clearly shown in various in vitro and animal models that IgG autoantibodies against BP180 are pathogenic [17][18][19][20][21][22] and correlate with disease activity [23][24][25][26] in BP patients. Most effects of anti-BP180 IgG are Fc receptor-mediated.…”

Value of BIOCHIP Technology in the Serological Diagnosis of Pemphigoid Gestationis

“…The BIOCHIP ® mosaic assay specifically detects IgG auto-antibodies bound to the respective auto-antigens. Circulating anti-BP230 IgG auto-antibodies were determined in parallel by 2 different ELISA systems, using an E. coli-expressed fragment of the C-terminal globular domain of BP230 (Euroimmun; BP230-C; amino acids 2326-2649) and both the recombinant N-and C-terminal fragments (MBL, Nagoya, Japan; BP230-N-C; amino acids 1-979 and 1869-2649) (12,13). All assays were performed according to the manufacturers' instructions.…”