CAJ | 학술논문

目的制备抗重组 GST的单克隆抗体 (mAb), 并用来纯化重组 GST融合蛋白。方法用含重组 GST融合蛋白基因的 pGEX4T -1质粒转化 E.coli BL21， IPTG诱导 GST融合蛋白表达，亲和层析和凝胶过滤法分离表达的重组 GST融合蛋白。以此蛋白作为抗原，免疫 Balb/c小鼠，按传统的杂交瘤技术制备 mAb。将抗 GST mAb经 Protein A纯化后，与 Sepharose 4B偶联。结果经 3次亚克隆后，获得两株分泌抗 GST载体特异性 mAb的杂交瘤。采用该 mAb对两种不同的 GST融合蛋白进行亲和层析纯化后，经 SDS-PAGE鉴定达到了商品化 Glutathione-Resin的亲和层析纯化效果。结论用抗 GST蛋白特异性 mAb亲和层析纯化融合蛋白是一种经济、实用的方法，且可用于 Glutathione-Resin亲和层析纯化后的二次纯化。
목적제비항중조 GST적단극륭항체 (mAb), 병용래순화중조 GST융합단백。방법용함중조 GST융합단백기인적 pGEX4T -1질립전화 E.coli BL21， IPTG유도 GST융합단백표체，친화층석화응효과려법분리표체적중조 GST융합단백。이차단백작위항원，면역 Balb/c소서，안전통적잡교류기술제비 mAb。장항 GST mAb경 Protein A순화후，여 Sepharose 4B우련。결과경 3차아극륭후，획득량주분비항 GST재체특이성 mAb적잡교류。채용해 mAb대량충불동적 GST융합단백진행친화층석순화후，경 SDS-PAGE감정체도료상품화 Glutathione-Resin적친화층석순화효과。결론용항 GST단백특이성 mAb친화층석순화융합단백시일충경제、실용적방법，차가용우 Glutathione-Resin친화층석순화후적이차순화。
Aim To prepare and characterize a monoclonal antibody against recombinant glutathione S-transferase(GST) for purifying GST fusion protein. Methods The GST-follistatin fusion protein was expressed by using a pGEX4T-1 expression vector in Escherichia coli BL21 and purified by glutathione-resin affinity column chromatography. Then female Balb/c mice were immunized with the GST-FS, The immunized splenocytes were fused with NS-1 hybridoma cells. Dreparation of the mAb was used by conventional hybridoma techniqal. The mAb purified by protein A, was culpled with Sepharose4B to purify further GST fusion protein by affinity chromatography. Results The SDS-PAGE showed that the GST fusion protein could be purified effctively by specific mAb affinity chromatography as same as by glutathione-resin affinity chromatography. Conclusion mAb affinity chromatography will be a ecnomical and useful method and it can be used for secondary purification of GST fusion protein following glutathione-resin affinity chromatography.