用单克隆抗体3F1亲和层析纯化rHuIFN-α2a的研究

Affinity purification of recombinant human interferon α2a with monoclonal antibody 3F1

采用淋巴细胞杂交瘤技术，制备了针对ｒＨｕＩＦＮα２ａ的单克隆抗体（ｍＡｂ）杂交瘤细胞系３Ｆ１，对ｍＡｂ３Ｆ１识别ｒＨｕＩＦＮα２ａ表位、中和活性等特点进行了研究，并与ＣｅｌＴｅｃｈ公司的ｍＡｂＮＫ２做了平行比较。发现ｍＡｂ３Ｆ１与ｍＡｂＮＫ２均具有中和ｒＨｕＩＦＮα２ａ活性的作用，但二者识别ｒＨｕＩＦＮα２ａ分子的表位不同，表明在ｒＨｕＩＦＮα２ａ分子上至少有２个中和性表位。进一步研究发现，ｍＡｂ３Ｆ１具有良好的用于亲和层析纯化ｒＨｕＩＦＮα２ａ的性能，每毫升湿胶可回收０．８ｍｇ１．０ｍｇｒＨｕＩＦＮα２ａ，纯度达９５％以上，生物学活性达２×１０１１ｕ／ｇ，且ｍＡｂ３Ｆ１有助于ｒＨｕＩＦＮα２ａ的复性。提示ｒＨｕＩＦＮα２ａ分子上的中和性表位，在与ｍＡｂ３Ｆ１的结合和解离过程中。

A hybridoma cell line 3F1 was raised to secrete monoclonal antibody with specificity to recombinant human interferon α2a. The features of 3F1 such as epitope recognition, neutralizing activity to rHuIFN α2a were compared in parallel with monoclonal antibody NK2 from CellTech. The results showed that both 3F1 and NK2 possess the activity of neutralizing rHuIFN α2a, although they recognize different epitopes on rHuIFN α2a, suggesting there are at least two neutralizing epitopes available on rHuIFN α2a. Further investigation demonstrated that 3F1 is ideal for affinity purification of rHuIFN α2a, resulting from the recovery rate of 0.8 mg￣1.0 mg of rHuIFN α2a per ml affinity gel with per ml affinity gel the purity of more than 95% and activity of 2×10 11 u/g, and beneficial to the renaturation of rHuIFN α2a. Taken together, it is concluded that the association and dissociation of neutralizing epitope of rHuIFN α2a during affinity purification are able to improve the renaturation of rHuIFN α2a.