融合蛋白ProteinA残留检测方法的建立  

Development of a method for determination of residual protein A of a kind of Fc fusion protein

作　　者：毕华[1] 范文红[1] 史新昌[1] 丁有学[1] 刘兰[1] 饶春明[1] 

机构地区：[1]中国食品药品检定研究院,北京100050

出　　处：《药物分析杂志》2014年第2期297-300,共4页Chinese Journal of Pharmaceutical Analysis

基　　金："国家重大新药创制"科技重大专项(2012ZX09304010)

摘　　要：目的:建立一种融合蛋白的Protein A(ProA)残留检测方法。方法:利用Cygnus公司的F400检测试剂盒,用酶联免疫吸附测定(ELISA)法进行残留量检测,通过基质干扰试验、校正标准曲线法和非校正标准曲线的对比及加标回收率试验考察基质的干扰。用校正标准曲线法对3批融合蛋白原液进行残余ProA检测,并对该方法进行精密性和准确性验证。结果:产品基质对试验检测有干扰,应用校正标准曲线法可以消除这种干扰。3批供试品原液经3次测定,残余ProA的平均值分别为(0.24±0.03)、(0.29±0.03)和(0.36±0.03)ng·mL-1,RSD均小于15%。1批原液经3次测定,回收率为(99.92±8.28)%。结论:已成功建立该融合蛋白的ProA残留量检测方法,重复性好,准确性高,可作为该融合蛋白ProA残留量的常规检测方法,也为融合蛋白及单抗类制品的ProA残留量检测提供借鉴。Objective： To develop a method for determination of Protein A （ProA） of a kind of fusion protein. Methods： The residual ProA was determined by ELISA using F400 kit of Cygnus Technologies.The matrix interference was inspected by interference test,comparison of calibration standard curve and non-calibration standard curve and recovery rate test.Three batches of bulks were determined by the calibration standard curve method,and the developed method was verified for precision and accuracy. Results： The matrix was proved to have an interference,and the interference could be removed by the calibration standard curve method.Three batches of bulks were determined for 3 times,and the result showed residual ProA mean values were （0.24±0.03）,（0.29±0.03） and （0.36±0.03） ng·mL^-1,,with a variation coefficient of less than 15%.One batch of bulk was determined for 3 times,and the result showed a recovery rate of （99.92 ± 8.28）%. Conclusion： A method for determination of residual ProA of a kind of fusion protein is successfully developed,which shows good reproducibility and high accuracy and may be used routinely,and supplies a reference for residual ProA determination of fusion protein and monoantibody products.

关 键 词：融合蛋白 ProA残留检测 酶联免疫吸附测定(ELISA) 校正标准曲线 验证 

分 类 号：R917[医药卫生—药物分析学]