摘要
目的对CHO-K1细胞簇集法检测百日咳毒素(pertussis toxin,PT)毒性进行优化及验证。方法以国家PT标准品为参考品,将PT标准品倍比稀释后与CHO-K1细胞混合培养,显微镜观察PT能引起CHO-K1细胞簇集的程度。对细胞浓度、孵育时间进行优化,并对方法的特异性、敏感度、重复性进行验证。结果方法的最佳细胞浓度为2.5×105个/ml,最佳孵育时间为24 h。只有PT活性成分能使CHO-K1细胞发生簇集,最低检测限为10 pg/ml;2名试验员6次检测结果表明,该方法重复性良好。结论 CHO-K1细胞簇集法可及时、准确地检测出生产过程中PT含量,适用于百日咳发酵过程监测。
Objective To optimize and verify the CHO-K1 cell cluster method for determination of toxicity of pertussis toxin. Methods National PT standard as a reference was diluted 2-fold serially with CHO-K1 cells. The cluster of CHOK1 cells caused by PT was observed by microscopy. The cell concentration and incubation time were optimized, and the method was verified for specificity, sensitivity and reproducibility. Results The optimal cell concentration and incubation time were 2. 5 × 10^5 cells / ml and 24 h respectively. Only the active ingredients of PT caused the cluster of CHO-K1 cells. The minimum detection limit of the method was 10 pg / ml. The six test results by two staff proved good reproducibility of the method. Conclusion CHO-K1 cell cluster method may determine the PT content during production timely and accurately, which is suitable for the monitoring the fermentation process of Bordetella pertussis.
出处
《中国生物制品学杂志》
CAS
CSCD
2015年第12期1344-1346,共3页
Chinese Journal of Biologicals
