DETECTION OF IGG AUTOANTIBODIES IN THE SERA OF PATIENTS WITH BULLOUS AND GESTATIONAL PEMPHIGOID - ELISA STUDIES UTILIZING A BACULOVIRUS-ENCODED FORM OF BULLOUS PEMPHIGOID ANTIGEN-2

Autoantibodies against the extracellular domain of bullous pemphigoid antigen 2 (BPAG2) are thought to play a key role in the pathogenesis o f bullous pemphigoid and their detection may thus be of diagnostic and prognostic value. The aim of this study was to develop a standardized enzyme-linked immunosorbent assay utilizing the baculovirus-derived p rotein BV13 (extracellular domain of BPAG2 devoid of 68 amino acids at the C terminus linked to glutathione-S-transferase and 6x His tag) to detect BPAG2-specific autoantibodies. For the enzyme-linked immunosor bent assay, nickel agarose affinity-purified BV13 protein was incubate d with sera from patients with bullous pemphigoid (n = 39), gestationa l pemphigoid (n = 10), and pemphigus vulgaris/pemphigus foliaceus (PV/ PF; n = 15), or normal human sera (NHS; n = 18). Nickel affinity-purif ied proteins from wild-type baculovirus-infected insect cells served a s a control. A positive enzyme-linked immunosorbent assay value was de fined as reactivity (ODBV13-ODWT) > mean reactivity + 1 SD of the nega tive control sera (PV/PF; NHS). Thirty-five of 39 bullous pemphigoid s era and 10 of 10 gestational pemphigoid sera were reactive to BPAG2 co mpared with none of 15 PV/PF sera and one of 18 NHS (sensitivity, 91.8 %; specificity, 97%). Of 16 BPAG2-reactive sera in the enzyme-linked i mmunosorbent assay, only six were BPAG2-reactive in the western blot, whereas 14 sera immunoprecipitated BPAG2 from extracts of epidermal ke ratinocytes. The enzyme-linked immunosorbent assay utilizing an eukary otic BPAG2 protein thus seems to be highly sensitive and specific in t he detection of BPAG2-specific antibodies and, hence, may be useful in the diagnosis of bullous autoimmune diseases, such as bullous pemphig oid and gestational pemphigoid.