重组葡激酶-水蛭素融合蛋白质控方法和质量标准的建立  

Development of methods and standards for quality control of recombinant staphylokinase-hirudin fusion protein

作　　者：丁有学[1] 韩春梅[1] 李响[1] 毕华[1] 史新昌[1] 饶春明[1] 

机构地区：[1]中国食品药品检定研究院重组药物室卫生部生物技术产品检定方法及其标准化重点实验室,北京100050

出　　处：《中国生物制品学杂志》2014年第8期1024-1028,1034,共6页Chinese Journal of Biologicals

基　　金：国家科技重大专项课题(2012ZX0930410)

摘　　要：目的建立注射用重组葡激酶(staphylokinase,SAK)-水蛭素(hirudin,HV)融合蛋白(SFH)的质控方法和质量标准。方法采用纤维蛋白平板溶圈(fibrin agarose plate assay,FAPA)法测定SFH的溶栓比活性,纤维蛋白凝块溶解法测定SFH的抗凝比活性;还原型SDS-PAGE测定SFH的相对分子质量;非还原SDS-PAGE和反相高效液相色谱(reversed-phase high-performance liquid chromatography,RP-HPLC)测定SFH的纯度;胰酶裂解后采用RP-HPLC法分析SFH的肽图;其他各项指标的检测按《中国药典》三部(2010版)规定进行。结果用建立的方法对SFH原液和成品进行检定,各项指标均符合《人用重组DNA制品质量控制技术指导原则》和《中国药典》三部(2010版)的要求。结论建立的质控方法和质量标准能够保证产品安全、有效、质量可控,可用于注射用SFH产品的常规检定。Objective To develop the methods and standards for quality control of recombinant staphylokinase-hirudin fusion protein （SFH） for injection. Methods SFH was determined for thrombolytic activity by fibrin agar plate assay （FAPA）, for anticoagulant activity by clot dissolution method, for relative molecular mass by reduced SDS-PAGE, and for purity by non-reduced SDS-PAGE and reversed-phase high-performance liquid chromatography （RP-HPLC）, then subjected to peptide mapping by RP-HPLC after digestion with trypsin. Other control tests were carried out according to the requirements in Chinese Pharmacopoiea （Volume III, 2010 edition）. Results Control tests were performed on the bulk and final product of SFH by the developed methods, and the result showed that all the indexes met the requirements in Chinese Pharmacopoiea （Volume III, 2010 edition）. Conclusion The developed methods and standards ensured the safety, efficiency and controllable quality of SFH, and might be used for the routine quality control of SFH.

关 键 词：重组葡激酶-水蛭素融合蛋白(SFH) 质量控制 

分 类 号：Q789[生物学—分子生物学]