以一步离子交换层析法制备人凝血因子Ⅷ/血管性血友病因子混合物的研究

Study on preparation of human coagulation factor Ⅷ/von Willebrand factor complex by one step ion exchange chromatography

目的研究一步离子交换层析法制备人凝血因子Ⅷ(factor Ⅷ,FⅧ)/血管性血友病因子(von Willebrand factor,vWF)混合物的最适层析条件。方法以含不同浓度氯化钠的缓冲液进行线性梯度洗脱来确定层析缓冲液的最适离子强度和pH值,同时研究上样体积对该法的影响。结果使用pH7.0的含250 mmol/L氯化钠的洗涤缓冲液和含400 mmol/L氯化钠的洗脱缓冲液进行离子交换层析,所得产物的FⅧ与vWF效价比约为1∶1,FⅧ回收率为76.51%。结论采用一步离子交换层析法制备FⅧ/vWF混合物,可使FⅧ与vWF效价比达到约1∶1。

Objective To study the optimal chromatography condition for preparing human coagulation factor Ⅷ(FⅧ)/ von Willebrand factor (vWF) complex by one step ion exchange chromatography (IEC). Methods The optimal ionic strength and pH value of chromatography buffer solution were identified by linear gradient elution using buffer solution containing different concentrations of sodium chloride. The influence of sample volume on IEC was also studied. Results When IEC was carried out using washing buffer containing 250 mmol/L sodium chloride and elution buffer containing 400 mmol/L sodium chloride at pH7.0, the activity ratio of FⅧ to vWF in obtained product was about 1∶1, and the recovery rate of FⅧ was 76.51%. Conclusion Using one step IEC to prepare FⅧ/vWF complex can achieve the activity ratio of FⅧ to vWF at about 1∶1.