HL60-IL6单核细胞活化反应法在注射用肝水解肽和注射用辅酶Ⅰ热原检测中的应用研究  

Application of HL60-IL6 Monocyte-activation Test to Pyrogen Testing of Heparolysate for Injection and Coenzyme Ⅰ for Injection

作　　者：王灿[1] 王鸣人 郑璐侠[1] 邵泓[1] 陈钢[1] Wang Can;Wang Mingren;Zhen Luxia;Shao Hong;Chen Gang(NMPA Key Laboratory for Quality Control of Therapeutic Monoclonal Antibodies,Shanghai Institute for Food and Drug Control,Shanghai 201203,China)

机构地区：[1]上海市食品药品检验所国家药品监督管理局治疗类单抗质量控制重点实验室

出　　处：《中国药师》2019年第12期2288-2291,共4页China Pharmacist

基　　金：上海市自然科学基金项目(编号:17ZR1426000)

摘　　要：目的:探讨HL60-IL6单核细胞活化反应法在注射用肝水解肽和注射用辅酶Ⅰ热原检测中的应用。方法:参照2015年版中国药典通则1193,确定注射用肝水解肽和注射用辅酶Ⅰ热原测定限值及最小有效稀释浓度。采用细胞增殖抑制试验、IL-6 ELISA试剂盒干扰试验和热原干扰试验分别探讨对HL60细胞生长、对IL-6和热原测定无影响的供试品浓度,进而确定适用于注射用肝水解肽和注射用辅酶Ⅰ体外热原测定的HL60-IL6方法。用该方法对1个厂家7批注射用肝水解肽、1个厂家6批注射用辅酶Ⅰ进行热原测定。结果:注射用肝水解肽和注射用辅酶Ⅰ最小有效稀释浓度分别为0.003 mg·ml^-1和0.0002 mg·ml^-1;对HL60细胞增殖无影响的最高浓度分别为1 mg·ml^-1和10 mg·ml^-1,对IL-6和热原测定无影响的最高浓度均为0.1 mg·ml^-1。根据以上实验结果确定了适合该方法的注射用肝水解肽和注射用辅酶Ⅰ浓度均为0.1 mg·ml^-1。用该方法对1个厂家7批注射用肝水解肽、1个厂家6批注射用辅酶Ⅰ供试品进行热原测定,结果均为阴性。结论:HL60-IL6方法适用于注射用肝水解肽和注射用辅酶Ⅰ热原测定。Objective:To explore the application of HL60-IL6 monocyte-activation test in the detection of pyrogen in heparolysate for injection and coenzymeⅠfor injection.Methods:According to the general rule 1193 in Chinese Pharmacopoeia(2015 edition),the limit of determination and the minimum effective dilution concentration of pyrogen in heparolysate for injection and coenzymeⅠfor injection were determined.The sample concentrations without effects on HL60 proliferation,pyrogen test and IL-6 test were determined by cell proliferation inhibition test,interference test of IL-6 ELISA kit and pyrogen interference test.According to the results,the in vitro pyrogen assay of heparolysate for injection and coenzymeⅠfor injection was established.And then,the pyrogen in 7 batches of heparolysate for injection from another manufacturer and 6 batches of coenzymeⅠfor injection from anther manufacturer were determined by the method.Results:The minimum effective dilution concentration of heparolysate for injection and coenzyme I for injection was 0.003 mg·ml^-1 and 0.0002 mg·ml^-1,respectively.The concentrations without effects on HL60 proliferation were 1 mg·ml^-1 and 10 mg·ml^-1,while those without effects on pyrogen test and IL-6 test were both 0.1 mg·ml^-1.As a results,the suitable concentration was 0.1 mg·ml^-1 of heparolysate for injection and coenzymeⅠfor injection,and the suitable monocyte-activation test of heparolysate for injection and coenzymeⅠfor injection was established.The pyrogen detection results were all negative.Conclusion:The method is suitable for the pyrogen detection of heparolysate for injection and coenzymeⅠfor injection.

关 键 词：单核细胞活化反应法 HL60-IL6 注射用肝水解肽 注射用辅酶Ⅰ 

分 类 号：R917[医药卫生—药物分析学]