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目的对汉坦病毒G2糖蛋白多表位抗原(multi-epitope antigen,MEA)进行原核表达和纯化,建立MEAELISA方法并用于肾综合征出血热(HFRS)患者血清特异性抗体的检测。方法利用pET原核表达系统表达MEA,并进行镍亲和纯化;利用Western blot方法对MEA进行免疫反应性鉴定,并建立MEA-ELISA方法,用于检测HFRS患者血清特异性抗体。结果原核表达系统E.coli BL21/pET32a-mea能表达分子质量单位为36.9ku的MEA蛋白,纯化后的蛋白浓度为0.15mg/ml,纯度>95%,Western blot显示目的蛋白能被HFRS患者血清识别。用建立的MEA-ELISA方法检测120例HFRS患者血清的阳性率为90.83%,检测60例健康人血清全部为阴性。结论重组表达蛋白MEA具有良好的免疫反应性,建立的MEA-ELISA方法可用于HFRS患者血清特异性抗体的检测。
Objective To express and purify the glycoprotein multi-epitope antigen (MEA) of Hantaviruses, and to establish the MEA ELISA method for the detection of serum specific antibodies in patients with hemorrhagic fever with renal syndrome (HFRS). Methods MEA was expressed by prokaryotic expression system of pET and purified by nickel affinity. The immunoreactivity of MEA was identified by Western blot and MEA-ELISA method was established for the detection of serum specific antibodies in HFRS patients. Results The prokaryotic expression system of E. coli BL21 / pET32a-mea expressed MEA protein with a molecular mass of 36.9ku. The purified protein was 0.15mg / ml with a purity of> 95%. Western blot showed that the target protein could be expressed in serum of HFRS patients Recognize. The positive rate of serum of 120 patients with HFRS detected by MEA-ELISA method was 90.83%. The serum of 60 healthy people were all negative. Conclusion The recombinant protein MEA has good immunoreactivity. The established MEA-ELISA method can be used to detect serum specific antibodies in HFRS patients.