摘要
目的构建人锌转运子ZnT8基因N末端、C末端和N-C融合区段的原核表达载体,诱导表达获得重组蛋白,并初步验证各抗原在1型糖尿病ZnT8自身抗体检测中的价值。方法应用RT-PCR方法调取目的基因,构建相应的原核表达质粒,转化大肠杆菌E.coli HB101,诱导表达获得纯化重组蛋白,并作为抗原包被酶联板,初步建立检测ZnT8自身抗体的ELISA方法 ,评价各基因片段在1型糖尿病诊断中的价值。结果获得了3种可被1型糖尿病患者血清识别的重组人ZnT8抗原区段,其中C末端区段ZnT8(268-369aa)与其它两个片段相比具有更高的检测敏感性和特异性,成为首选的抗原区段。结论所选重组人ZnT8(268-369aa)抗原区段具有良好的抗原性,可作为1型糖尿病患者辅助诊断试剂的候选抗原。
Objective To obtain different fragments of human zine transporter ZnT8 by construction and inducing the prokaryotic expression vector of ZnT8,and to evaluate the diagnostic application for type 1 diabetes mellitus of the recombination human ZnT8.Methods The coding gene of the ZnT8 was obtained by RT-PCR.The corresponding prokaryotic expression vectors pBVIL1/ZnT8 were constructed and transformed into E.coli HB101 to inducing the expression of the recombination human different fragments of human ZnT8.Using the two antigen fragments as the coating antigens,the enzyme-linked immunosorbent assay（ELISA） was established for the detection of the ZnT8 autoantibody.Results The obtained two fragments of human ZnT8 could react with the serum of type 1 diabetes mellitus patients.The sensitivity and the specificity of the ZnT8（268-369） fragment were higher than that of the other fragments.Thus,the ZnT8（268-369aa） fragment became to the preferred antigen.Conclusion Because of the favourable antigenicity of the selected recombination human ZnT8（268-369aa）,the ZnT8（268-369aa） could be the candidate antigen for developing the diagnostic reagent for type 1 diabetes mellitus.
出处
《中国实验诊断学》
北大核心
2011年第1期102-106,共5页
Chinese Journal of Laboratory Diagnosis
