Envoplakin and Periplakin, the Paraneoplastic Pemphigus Antigens, are also Recognized by Pemphigus Foliaceus Autoantibodies Shideh Kazerounian, Mỹ G.

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Envoplakin and Periplakin, the Paraneoplastic Pemphigus Antigens, are also Recognized by Pemphigus Foliaceus Autoantibodies Shideh Kazerounian, Mỹ G. Mahoney, Jouni Uitto, Sirpa Aho Journal of Investigative Dermatology Volume 115, Issue 3, Pages 505-507 (September 2000) DOI: 10.1046/j.1523-1747.2000.00088-2.x Copyright © 2000 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 1 Periplakin and envoplakin linker domains are recognized on the western blot analysis by PNP and PF, but not by PV, patient sera. Recombinant GST-fusion proteins composed of conserved linker region within the C-terminal domains of periplakin (PP) and envoplakin (EV) (1 μg per lane) were separated on 10% SDS–PAGE. After transfer, filters were blocked with 1% BSA, 5% dry milk in PBS, and incubated overnight at 4°C with sera (1:1000) indicated. Bound antibodies were visualized with peroxidase conjugated anti-human IgG and the Renaissance Chemiluminescence Reagent (NEN Life Science Products). Sera from six PF patients (A), four PV patients (B), and two PNP patients (C ) were analyzed. Arrows on the left side of each panel indicate the position of periplakin–GST fusion protein, and the arrows on the right side of each panel point to the envoplakin–GST fusion protein. Journal of Investigative Dermatology 2000 115, 505-507DOI: (10.1046/j.1523-1747.2000.00088-2.x) Copyright © 2000 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 2 The envoplakin linker ELISA demonstrates autoanti-bodies from PNP and PF patient sera. Microtiter plate wells were coated with envoplakin linker–GST fusion protein and recombinant GST (400 ng per 50 μl) in 0.1 M bicarbonate buffer, pH 9.6, at 4°C overnight, washed with PBS, and blocked with 100 μl of PBS containing 1% BSA. Patient sera were diluted (1 : 800), applied in triplicates (50 μl per well), and incubated at 4°C overnight, washed and then incubated with 1000-fold diluted alkaline phosphatase-conjugated anti-human antibody for 1 h. The mean of triplicate assays with paraneoplastic pemphigus sera (PNP), pemphigus foliaceus sera (PF+ PF982, PF1396, PF1410, PF3009), (PF–, PF 1738, PF2099) and pemphigus vulgaris sera (PV) are blotted as optical density at 405 nm. As negative control, the results obtained with 1:800 dilution of normal sera, as well as those obtained by omitting either primary or secondary antibody, are shown (N). The letters within the circles refer to the individual sera identified in Figure 1. Journal of Investigative Dermatology 2000 115, 505-507DOI: (10.1046/j.1523-1747.2000.00088-2.x) Copyright © 2000 The Society for Investigative Dermatology, Inc Terms and Conditions