一种抗人T淋巴细胞免疫球蛋白淋巴细胞毒效价检测方法的建立  

Development of a method for determination of anti-human T lymphocyte immunoglobulin lymphocytotoxic titer

作　　者：杜天飞 容新宗 杨盛理 张勇侠[1] 武志强 李春阳[1] 刘兰军[1] DU Tian-fei;RONG Xin-zong;YANG Sheng-li;ZHANG Yong-xia;WU Zhi-qiang;LI Chun-yang;LIU Lan-jun(Chengdu Institute of Biological Products Co.,Ltd,Center for Sichuan Province Vaccine Engineering Research,Chengdu 610023,Sichuan Province,China)

机构地区：[1]成都生物制品研究所有限责任公司四川省疫苗工程技术研究中心,四川成都610023

出　　处：《中国生物制品学杂志》2022年第12期1500-1505,共6页Chinese Journal of Biologicals

摘　　要：目的建立基于人白血病T细胞株Jurkat的抗人T淋巴细胞免疫球蛋白(anti-human T lymphocyte immunoglobulin,ALG)淋巴细胞毒效价定量检测方法,并进行验证。方法采用人Jurkat细胞作为靶细胞,抗人T细胞兔免疫球蛋白(商品名:Grafalon)作为待检样品,建立其淋巴细胞毒效价定量检测方法。优化反应体系,考察细胞培养密度、细胞代次对待检样品效价的影响,并对方法的精密度进行验证。结果优化的反应体系组成为50μL待检样品+50μL补体(1∶5稀释)+50μL细胞(5×10^(6)个/mL);细胞传代密度为2×10^(5)个/mL传代培养72 h或3×10^(5)个/mL传代培养48 h用于效价测定最佳;当细胞生长状态较好且一致时,从P10到P25代次的细胞效价测定结果较一致。采用该方法板内重复测定待检样品效价,变异系数(CV)为6.32%,重复性良好;不同代次Jurkat细胞测定待检样品效价6次,CV为4.15%,中间精密度良好。结论建立了一种便捷、精确的测定ALG淋巴细胞毒效价的定量检测方法,为ALG效价测定提供了一种客观、准确的评价手段。Objective To develop and verify a quantitative assay for determination of anti-human T lymphocyte immunoglobulin(ALG)lymphocytotoxic titer based on human leukemia T cell line Jurkat.Methods Using human Jurkat cells as target cells and anti-human T cell rabbit immunoglobulin(trade name:Grafalon)as the sample to be tested,a method for quantitative determination of the lymphocytotoxic titer was developed,which was optimized for the reaction system,investigated for the effects of culture density and generation of cells on the titer of the sample,and verified for the precision.Results The optimized reaction system consisted of 50μL sample+50μL complement(1∶5 diluted)+50μL cells(5×10^(6)cells/mL).Cell subculture density of 2×10^(5)cells/mL for 72 h subculture or 3×10^(5)cells/ml for 48 h subculture was optimal for determination of titer,and the determination results from P10 to P25 generations were consistent when the state of cell growth was good and uniform.CV value of intraplate repetitive determination of the titer of sample was 6.32%,showing a good repeatability.Different generations of Jurkat cells were used to determine the titer of sample for 6 times,with a CV value of 4.15%and a good intermediate precision.Conclusion A convenient and accurate quantitative assay for determination of ALG lymphocytotoxic titer was developed,which provided an objective and accurate evaluation method for the determination of ALG titer.

关 键 词：抗人T淋巴细胞免疫球蛋白 JURKAT细胞 淋巴细胞毒活性 

分 类 号：R392-33[医药卫生—免疫学]