可溶性肝抗原的克隆、表达及初步临床应用

为克隆、表达可溶性肝抗原 (SLA ) ,本文采用RT PCR技术从人肝组织基因中扩增SLA全长cDNA ,经双酶切插入载体PQE 30并在大肠杆菌M15中表达。对表达载体PQE 30 /SLA中的SLA进行序列分析 ,表达产物用SDS PAGE、免疫印迹方法鉴定并初步临床应用。结果显示SLAcDNA序列与报道一致 ,表达产物在相对分子质量略大于 4 7× 10 4处有一条明显的蛋白带 ,该蛋白带能特异性与抗SLA阳性血清反应。表明构建的表达载体PQE 30 /SLA能表达具有抗原活性的可溶性肝抗原。以SLA融合蛋白为抗原检测 18例临床诊断为自身免疫肝炎患者 ,4例SLA抗体阳性。重组SLA融合蛋白的制备为自身免疫性肝炎的诊断及其发病机制研究提供物质基础

Cloning and Expression of the Soluble Liver Antigen and Its Clinical Trial

To clone and express the soluble liver antigen(SLA) by using the RT PCR technique and other related methods,the SLA cDNA was obtained by amplification of the full length of cDNA from human liver tissues Then it was inserted into fusion expression vector PQE 30 site of BamH I and Hind III and expressed in E coli M15 The expressed products were identified by SDS PAGE and Western blot Results on SDS PAGE analysis explored a very clear band at site of molecular weight of more than 47 5 kD with specific reaction to the anti SLA antibody.These results indicate that the vector expressed PQE 30/SLA has the ability to express SLA with antigenic activity The SLA fusion protein was used as antigen for a preliminary clinical trial in 18 cases of autoimmune hepatitis with 4 cases showing positive SLA reaction It concludes that the cloning and expression of SLA would provide a useful material for the diagnosis and scientific research on autoimmune hepatitis