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目的：克隆并表达寻常性天疱疮抗原（PVA，即桥粒芯糖蛋白-3）的EC1-2和EC3-4表位，用于通过血清学方法特异性诊断天疱疮和了解PVA的表位与抗PVA抗体间的联系。方法：从角朊细胞抽提RNA，逆转录合成cDNA,扩增目的基因EC1-2和EC3-4后与质粒载体PGEX－4T-1连接，电转导入大肠杆菌BI21，经IPTG诱导后表达EC1-2和EC3-4融合蛋白，SDS-PAGE电泳后转移至硝酸纤维素膜，免疫印迹法检测抗PVA抗体。结果：经限制性内切酶分析和DNA序列测定，克隆的EC1－2、EC3-4基因与PC／CGENE登录的序列一致；表达的重组蛋白仅与寻常性天疱疮血清反应，不与大疱性类天疱疮、系统性红斑狼疮以及正常人血清反应。结论：该重组蛋白具有高度抗原特异性，此项研究为通过血清学方法诊断寻常性天疱疮和鉴别诊断其它大疱性皮肤病提供了方法，为进一步研究粘附分子与天疱疮发病的时间关系奠定了基础。
AIM: To clone and express the EC1-2 and EC3-4 epitopes of the pemphigus antigen (PVA, desmoglep-3) for the specific diagnosis of pemphigus by serological methods and for the understanding of PVA Relationship between epitopes and anti-PVA antibodies. METHODS: RNA was extracted from keratinocytes, cDNA was reverse transcribed, and the target genes EC1-2 and EC3-4 were amplified and ligated into plasmid vector PGEX-4T-1. The recombinant plasmid was transformed into E. coli BI21 and induced by IPTG to express EC1- 2 and EC3-4 fusion protein, after SDS-PAGE electrophoresis transferred to nitrocellulose membrane, Western blot detection of anti-PVA antibody. Results: Restriction endonuclease analysis and DNA sequence analysis showed that the cloned EC1-2 and EC3-4 genes were identical with the sequences registered by PC / CGENE. The expressed recombinant protein only reacted with the serum of pemphigus vulgaris, Bullous pemphigoid, systemic lupus erythematosus, and normal human serum. CONCLUSION: The recombinant protein is highly antigen specific. This study provides a method for the diagnosis of pemphigus vulgaris by serological methods and the differential diagnosis of other bullous dermatoses. In order to further study the pathogenesis of adhesion molecules and pemphigus Time laid the foundation for the relationship.