重组人脑髓鞘碱性蛋白及其抗体的研究

将EcoR1和SalⅠ酶切的人脑髓鞘碱性蛋白（MBP）基因cDNA克隆片段与表达载体pGEX-5T重组后转化大肠杆菌，经筛选，增殖和IPTG诱导，阳性克隆SDS－PAGE结果证实表达一条特异42KDa区带，Western印记杂交证实该区带具MBP抗原特异性，免疫斑点杂交和ELISA检测表达产量占菌体可溶性蛋白含量6％，达到414.6mg/L菌液，将含可溶性MBP蛋白的菌液后SDS－PAGE分离纯化得重组MBP抗原，对新西兰兔进行背部皮下多点注射，5次免疫后以琼脂板免疫双扩法检测抗效价达1：16，并通过免疫斑点杂交和Western印亦杂交证实证抗体具抗MBP特异性。

Recombinant Human Brain Myelin Basic Protein and Its Antibody

We constructed the expression vector by inserting 21.5 KDa MBP human brain full-length cDNA coding sequence digested with restriction enzyme EcoR I and Sal I into downstream of pGEX-5T expression vector. The recombinant vector p5TMP was transformed into E. coli and the positive clonies were selected and incubated in LB medium induced by IPTG (isopropyl- -D-thiogalactoside). A new polypeptide band with apparent molecular weight 42 KDa was detected in transformed cell lysates by SDS-PAGE. Western blotting analysis confirmed that this fusion protein reacted specifically with antibodies to MBP, the expression level of MBP was about 414.6 mg/L medium estimated by immuno-dot blot, ELISA and absorbance scanning. Newzealand rabbits were immunized by subcutaneous injection of the purified recombinant MBP. The titer was obtained at 1:16 after 5 injections. The specificity of the antibody to MBP was confirmed by immuno-blot and Western blotting.