重组粉尘螨Ⅱ类变应原的原核表达及纯化

目的表达并纯化粉尘螨2类变应原Der f 2蛋白,检测其变应原性。方法用1mmol/L的IPTG诱导含重组质粒pET30a-Der f2的大肠埃希菌BL21(DE3),SDS-PAGE电泳检测并纯化目的蛋白,ELISA检测其与尘螨哮喘患者血清IgE的结合活性。结果 SDS-PAGE电泳检测重组质粒pET30a-Der f 2表达产物,其分子质量单位为15ku,与预期大小相符。ELISA检测纯化的Der f 2蛋白可与尘螨哮喘患者血清IgE反应。结论成功获得并纯化了Der f2重组蛋白,该蛋白具有变应原性,为进行尘螨过敏性疾病的变应原特异性免疫治疗的动物实验奠定了物质基础。

Prokaryotic expression and purification of recombinant products of group 2 house dust mite allergen

Objectives To express and purify the cDNA of the group 2 allergen of Dermatophagoides farinae(Der f 2) and to determine the allergic activity of the resulting recombinant protein.Methods The Der f 2 gene was inserted into the expression vector pET-30a(+) and transformed into competent E.coli BL21(DE3).The product induced by IPTG was analyzed with SDS-PAGE.The ability of Ig E to bind with sera from patients with an allergy to house dust mites was detected.Results SDS-PAGE testing showed that the amplified product with recombinant vector pET-30a-Der f 2 had a molecular mass of 15ku.This is consistent with the expected value.ELISA testing showed that the protein bound with the serum Ig E from patients with asthma.Conclusion Der f 2 has been successfully expressed in E.coli and the purified recombinant protein has good allergenicity,providing a basis for animal testing of ASIT to treat allergy to house dust mites.