用于人凝血因子Ⅷ生产的DEAE 650M层析凝胶使用寿命的评价

目的评价用于人凝血因子Ⅷ生产的DEAE 650M层析凝胶的使用寿命。方法在人凝血因子Ⅷ生产用DEAE 650M凝胶层析系统基础上,将柱体积缩小353倍,建立DEAE 650M凝胶层析柱缩小模型。采用该模型连续层析10批次人凝血因子Ⅷ样品,每次层析前均用1 mol/mL的NaCl上样,检测理论塔板数和对称系数,以确认柱效;同时分析样品比活性、层析收率、层析纯化倍数、层析上样比例。结果 DEAE 650M凝胶层析缩小模型每次层析人凝血因子Ⅷ样品前,理论塔板数均> 2 000,对称系数均在1. 0~1. 4范围内;纯化后样品比活性均高于20 IU/mg,层析收率均高于40%,层析纯化倍数均大于20倍,层析上样比例均大于25 IU/mL。结论 DEAE 650M凝胶用于人凝血因子Ⅷ生产至少可循环使用10次。

Evaluation on service life of DEAE 650M chromatographic gel for production of human coagulation factor Ⅷ

Objective To evaluate the service life of DEAE 650 M chromatographic gel for production of human coagulation factor Ⅷ(FⅧ). Methods The volume of chromatographic column for production of FⅧ by using DEAE 650 M gel was reduced by 353 folds to establish a miniature model,by which ten consecutive batches of F Ⅷ samples were purified. Sodium chloride at a concentration of 1 mol/mL was laoded before each chromatography. The theoretical plate number and symmetry coefficient were calculated the confirm the efficacy of chromatographic column. Meanwhile,the specific activity,recovery,purification fold and loading proportion of samples were analyzed. Results All the theoretical plate numbers of ten consecutive batches of FⅧ samples purified by DEAE 650 M gel chromatography were more than 2 000,while the symmetry coefficients were 1. 0 ~ 1. 4. The specific activities of purified samples were more than 20 IU/mg while the recoveries were more than 40%,the purification folds were more than 20,and the loading proportions were more than 25 IU/mL. Conclusion DEAE 650 M gel may be used for at least 10 times for production of FⅧ.