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目的:建立对重组人血管内皮生长因子抑制剂进行异天门冬氨酸(Isoasp)含量检测的方法。方法:利用ISOQUANT异天门冬氨酸检测试剂盒并结合HPLC的方法检测重组人血管内皮生长因子抑制剂原液和成品中的Isoasp含量,并对该方法进行加标回收率试验、精密度和准确性的验证。结果:40、25、6.25 pmol·L-1的3种浓度对照品的加标回收率均在80%~120%的范围内。对1批原液和1批成品的各3个样品分别进行测定,原液和成品中每摩尔蛋白的Isoasp含量平均值分别为(0.066±0.010)、(0.093±0.008)mol,RSD分别为15.2%和8.6%,原液和成品中各1个样品的25 pmol·L-1对照品的加标回收率分别为90.2%和108.1%。结论:经方法学验证,本文所建立的方法可作为该产品的Isoasp含量的常规检测方法,也为融合蛋白及单抗类制品的Isoasp含量检测提供借鉴。
Objective: To establish a method for the determination of Isoasp in recombinant human vascular endothelial growth factor inhibitor. Methods: The isoquat content in the recombinant human VEGF-A and its finished product was detected by ISOQUANT aspartate test kit and HPLC method. The precision and accuracy of the method were tested by spiked recoveries Verification. Results: The spiked recoveries of the three concentrations of 40, 25 and 6.25 pmol·L-1 were all in the range of 80% -120%. The isoasp content of the original solution and the finished product was (0.066 ± 0.010), (0.093 ± 0.008) mol, respectively, and the RSDs were 15.2% and The spiked recoveries of 8.6%, 25 pmol·L-1 reference product in each of the original solution and finished product were 90.2% and 108.1%, respectively. Conclusion: The method established in this paper can be used as a routine method for the determination of Isoasp content of the product, as well as for Isoasp content detection of the fusion protein and monoclonal antibody products.