细菌内毒素重组C因子检测方法的验证

目的验证细菌内毒素重组C因子检测法。方法根据《中国药典》四部(2015版)9101药品质量分析方法验证指导原则的规定,对细菌内毒素重组C因子检测法(Lonza和Hyglos两种试剂盒)的线性范围、准确度、精密性、专属性、检测限及耐用性进行验证,并与动态显色法进行比较。结果 2种方法在内毒素标准品为0. 005～5 EU/mL范围内与logδRFU均呈良好的线性关系,r均> 0. 980;高、中、低浓度内毒素标准品2种方法检测回收率的平均值差异均无统计学意义(P> 0. 05);精密度各有优劣,总体相当;重组C因子不与β-葡聚糖发生反应,动态显色法检测β-葡聚糖浓度为0. 55 EU/mL;检测限均为0. 005 EU/mL;斜率和实测值不受增益值改变的影响,截距和增益值呈正相关。结论重组C因子法具有良好的准确度、精密性、专属性及耐用性,可应用于生物制品的细菌内毒素检查。

Validation of a determination method for recombinant factor C of bacterial endotoxin

Objective To validate a determination method for recombinant factor C of bacterial endotoxin.Methods According to the<9101>Guidelines for the Verification of Pharmaceutical Quality Analysis Methods in Chinese Pharmacopoeia(VolumeⅣ,2015 edition),recombinant factor C test(Lonza and Hyglos kits)was validated for linear range,accuracy,precision,specificity,detection limit and durability,and the results were compared with those of dynamic chromogenic test.Results By both recombinant factor C and dynamic chromogenic tests,the standard endotoxin content at a range of 0.005~5 EU/mL showed good linear relationship to the logδRFU,with r values of more than 0.980.No significant differences were observed in the mean recovery rates of samples at high,moderate and low concentrations determined by the two methods(P>0.05).However,the precisions of the two methods were equivalent in general.Recombinant factor C showed no reaction withβ-glucan,while theβ-glucan concentration determined by dynamic chromogenic test was 0.55 EU/mL.Both the detection limits of the two methods were 0.005 EU/mL.The slope and measured value were not influenced by the change of gain,while the intercept was positively related to the gain.Conclusion Recombinant factor C test showed high accuracy,precision,specificity and durability,which might be used for the determination of bacterial endotoxin in biologics.