人凝血因子Ⅷ活性生物检定统计法(显色法)的建立

Development of biometric verification method (chromogenic method) for determination of biological activity of human coagulation factor Ⅷ

目的依据《欧洲药典》8.0版规定的生物统计检定法原理,建立检测人凝血因子Ⅷ（FⅧ）活性的显色法。方法采用斜率比模型中随机区组设计显色法实验,进行数据统计分析及结果计算。以FⅧ国际标准品及FⅧ原液、成品为测试样品,确定其线性范围,并验证其重复性、中间精密度、专属性;用《中国药典》三部（2010版）规定的一期法和建立的方法进行比较。结果当FⅧ效价在0-17.85 IU/L范围内时,标准品和样品的线性相关系数（r）均大于0.99;重复性相对标准偏差（RSD）为2.1%;2位实验人员测定结果经t检验,差异无统计学意义（P〉0.05）;该方法置信限范围为80%-120%。两种方法检测结果差异无统计学意义（P〉0.05）。结论该方法操作简便、快速,测定结果稳定、准确,检测水平符合《欧洲药典》要求,可用于FⅧ活性检测。

Objective To develop a chromogenic method for determination of biological activity of human coagulation factor Ⅷ（FⅧ）based on the principle of biometric verification method in European Pharmacopoeia 8. 0. Methods The procedure of chromogenic method was designed by randomized block in slope ratio model, while the data were analyzed statistically, and the result was calculated. The method was determined for linear range by using international standard,bulk and final product of F Ⅷ, and verified for reproducibility, intermediate precision and specificity. The determination result by the developed method was compared with that by one phase method in Chinese Pharmacopoeia（Volume Ⅲ, 2010edition）. Results Both the linear correlation coefficients（r values）of the developed method for determination of standard and samples at F Ⅷ titers of 0 - 17. 85 IU / L were more than 0. 99, while the relative standard deviation（RSD） in reproducibility test was 2. 1%. The determination results by two staff were analyzed by t test and showed no significant difference（P〉0. 05）. The confidence limit of the method was 80% - 120%. The determination results by the developed method showed no significant difference with that by one phase method in Chinese Pharmacopoeia（Volume Ⅲ, 2010edition）（P〉0. 05）. Conclusion The developed method was simple, rapid, stable and accuracy, which met the requirements in European Pharmacopoeia 8. 0 and might be used for the determination of biological activity of FⅧ.