Wu Tie, Bi Jingxiu, Huang Yongdong, Zhang Yan, Sun Lijing, Sun Chunbao, Su Zhiguo
Civil & Environmental Engineering School, University of Science and Technology ofBeijing, Beijing 100083, China.
Sheng Wu Gong Cheng Xue Bao. 2008 Jul;24(7):1279-84. doi: 10.1016/s1872-2075(08)60061-8.
The low recovery of pertussis toxin (PT) and the low resolving efficiency of chromatography, due to the instability of PT in low salt condition, are the main challenges for its purification. We aplied 2 mol/L urea to prevent the aggregation and disassociation of PT during the purification by ion-exchange chromatography (IEC) and gel filtration chromatography (GFC). The effect of urea on the purification of PT was studied by ELISA assay and non-reduced SDS-PAGE. The activity recoveries of PT and filamentous hemagglutinin (FHA) in IEC and GFC, the resolution efficiency in GFC and the purities of PT and FHA were improved obviously by adding 2 mol/L urea in the mobile phase. The results highlight the potential application of urea in the acellular pertussis vaccine (APV) manufacture procedure.
由于百日咳毒素（PT）在低盐条件下不稳定，导致其回收率低以及色谱分离效率低，这是其纯化过程中的主要挑战。我们应用2 mol/L尿素来防止PT在离子交换色谱（IEC）和凝胶过滤色谱（GFC）纯化过程中的聚集和解离。通过ELISA测定和非还原SDS-PAGE研究了尿素对PT纯化的影响。在流动相中添加2 mol/L尿素后，IEC和GFC中PT和丝状血凝素（FHA）的活性回收率、GFC中的分离效率以及PT和FHA的纯度均得到明显提高。结果突出了尿素在无细胞百日咳疫苗（APV）生产过程中的潜在应用。
