兔抗人胸腺/淋巴细胞免疫球蛋白药代动力学检测方法的建立  

Establishment of a pharmacokinetic assay for rabbit anti-human thymus/lymphocyte immunoglobulin

作　　者：邹浩勇 余泽琼 耿鹏 殷文曲 王吟 周小璐 聂希霖 Zou Haoyong;Yu Zeqiong;Geng Peng;Yin Wenqu;Wang Yin;Zhou Xiaolu;Nie Xilin(Research Development Department,Wuhan ZhongshengYujin Biomedical Co.,Ltd.,Wuhan 430207,China;Quality Control Department,Wuhan ZhongshengYujin Biomedical Co.,Ltd.,Wuhan 430207,China;Wuhan Institute of Biological Products Co.,Ltd.,Wuhan 430207,China)

机构地区：[1]武汉中生毓晋生物医药有限责任公司科研开发部,武汉430207 [2]武汉中生毓晋生物医药有限责任公司质量控制室,武汉430207 [3]武汉生物制品研究所有限责任公司,武汉430207

出　　处：《国际生物制品学杂志》2024年第5期285-291,共7页International Journal of Biologicals

摘　　要：目的建立兔抗人胸腺/淋巴细胞免疫球蛋白(rabbit anti-human thymus/lymphocyte immunoglobulin, r-ATG)药代动力学检测方法并验证, 用于检测临床用药后患者血清中r-ATG的浓度。方法采用鼠抗兔IgG包被酶标板, 辣根过氧化物酶标记的羊抗兔IgG作为二抗, 建立双抗体夹心ELISA检测人血清中r-ATG的含量, 并对该方法进行线性范围、平行性、重复性、中间精密度、准确性、专属性及不同时间参考品标准曲线浓度的回收率稳定性验证。用建立的方法对5例患者的临床血清样本(血样)进行检测并进行药代动力学分析。结果以0.25 μg/ml为起始浓度, 稀释倍数在21~29之间的参考品浓度与吸光度值呈S曲线, 决定系数(R^(2))>0.99, 回收率在90%~110%;参考品与3份血样的线性方程斜率的变异系数(coefficient of variation, CV)<10%, R^(2)>0.99, R^(2)的CV<5%, 平行性较好;重复性和中间精密度CV<15%, 回收率均在90%~110%;准确性验证结果平均回收率均在80%~120%;注射药物的患者血清与健康人血清以及未注射药物的患者血清之间的差异有统计学意义, t值分别为32.607、21.949, P<0.01;参考品在2~8 ℃放置16个月时标准曲线各浓度的回收率均值均在90%~110%, 且各浓度不同时间点回收率相对标准偏差<15%;5例患者均在给药第5天血药浓度达到最高值, 平均半衰期为(6.5±5.3) d。结论建立的方法具有良好的稀释线性和样本间的平行性、重复性、中间精密度、准确性、专属性, 且参考品标准曲线各浓度在不同时间点的回收率稳定性良好, 可以用于r-ATG临床用药后血清浓度的检测。Objective To establish and verify a pharmacokinetic detection method for rabbit anti-human thymus/lymphocyte immunoglobulin(r-ATG),for detection of the total concentration of r-ATG in the sera of patients after clinical medication.Methods Mouse anti-rabbit IgG was used to coat the ELISA plate,and horseradish peroxidase-labeled goat anti-rabbit IgG was used as the secondary antibody to establish a double-antibody sandwich ELISA method for the detection of r-ATG content in human serum.The linear range,parallelism,repeatability,intermediate precision,accuracy,specificity,and stability of recovery rates for standard curve concentrations of reference materials at different time points of the method were verified.Clinical blood samples of 5 patients were detected and pharmacokinetic analysis was carried out by the established method.Results When the initial concentration was 0.25μg/ml and the dilution times were between 21-29,the reference concentration and the absorbance value showed an S curve,the coefficient of determination(R^(2))was>0.99,and the recovery rate was 90%-110%.The coefficient of variation(CV)of slope of linear equation between reference sample and three blood samples was<10%,with R^(2)>0.99,CV of R^(2)<5%,showing good parallelism.The repeatability CV and intermediate precision CV were<15%,and the recovery rates were all 90%-110%.The average recovery rates of accuracy verification results were 80%-120%.There was statistically significant difference between the sera of the injected patients,healthy people,and non-injected patients,the t-values were 32.607 and 21.949,respectively,with P<0.01.The average recovery of each concentration in the standard curve was 90%-110%when the reference was stored at 2-8℃for 16 months,and the relative standard deviation of the recovery at different time points of each concentration was less than 15%.The serum concentration of r-ATG reached the highest value on the 5th day after administration,with an average half-life time of(6.5±5.3)d.Conclusion The established method

关 键 词：免疫球蛋白类 兔抗人胸腺/淋巴细胞免疫球蛋白 药代动力学 酶联免疫吸附试验 

分 类 号：R-392[医药卫生]