利用自动发酵罐培养毕赤酵母,甲醇诱导其分泌表达重组猪IFN-α(r Po IFN-α);经硫酸铵沉淀、G-25琼脂糖柱脱盐处理及阴离子交换柱和分子筛层析纯化。以纯化r Po IFN-α蛋白作为抗原免疫6周龄BALB/c鼠3次后,取小鼠脾细胞与SP2/0骨髓瘤细胞进行融合,经间接ELISA筛选和3~5次亚克隆,共获得5株均能稳定分泌抗r Po IFN-α单克隆抗体的杂交瘤细胞株。抗体亚类鉴定、特异性和叠加试验结果表明,这5株抗体均属于Ig G1亚类,能与r Po IFN-α产生特异性反应,其抗原结合位点相同或相近。该研究有助于进一步推动r Po IFN-α单克隆抗体在猪免疫学以及猪疫病诊断中的应用。
【英文摘要】
By the fermentation reactor and Pichia pastoris expression system,the expression of recombinant porcine interfer-α(r Po IFN-α) was induced with methanol. The r Po IFN-α was purified by ammonium sulfate precipitation,Q-Sepharose fast flow chromatography and gel filtration in turn. 6-week-old BALB / c mice were immunized for three times with purified r Po IFN-α,then mouse myeloma cells and the spleen cells of the BALB / c mice were fused. Five hybridoma clones secreting antibody against r Po IFN-α were obtain...
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