抗人T细胞猪免疫球蛋白药代动力学检测方法的建立、验证及应用  

Establishment,validation and application of a pharmacokinetic assay for porcine anti-human T cell immunoglobulin

作　　者：殷文曲 余泽琼 张智 徐玉娟 宋桂芝 周小璐 邹浩勇 Yin Wenqu;Yu Zeqiong;Zhang Zhi;Xu Yujuan;Song Guizhi;Zhou Xiaolu;Zou Haoyong(Research Development Department,Wuhan Zhongsheng Yujin Biomedical Co.,Ltd.,Wuhan 430207,China;Quality Control Department,Wuhan Zhongsheng Yujin Biomedical 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

出　　处：《国际生物制品学杂志》2022年第6期320-324,共5页International Journal of Biologicals

摘　　要：目的建立抗人T细胞猪免疫球蛋白(porcine anti-human T cell immunoglobulin,p-ATG)药代动力学检测方法并验证,以检测用药后患者血清总p-ATG浓度。方法采用兔抗猪IgG F(ab′)2包被酶标板、辣根过氧化物酶标记的羊抗猪IgG作为二抗,建立人血清p-ATG含量的双抗体夹心ELISA,并验证该方法的标准曲线拟合范围、平行性、精密度、准确度、专属性。用建立的方法对13例患者的不同用药剂量的临床血样进行检测并分析药代动力学。结果从0.44μg/ml起始浓度稀释21~27倍的参考品浓度与吸光度值之间呈现S曲线,决定系数(R2)大于0.99,回收率为89.91%~112.96%;平行性样品斜率与参考品斜率变异系数(coefficient of variation,CV)绝对值<10%,R2>0.99,R2的CV<5%;板间、板内曲线CV<10%;重复性CV<10%,回收率为94.23%~101.80%;加标回收率均在99.87%~104.94%之间;输注p-ATG后患者血清与健康人血清以及输注前患者血清p-ATG浓度间差异有统计学意义(F=1.48,P<0.05)。20、25、30 mg剂量用药患者的血样中,p-ATG半衰期分别为20.6、16.6、16.2 d。结论建立的p-ATG药代动力学检测方法具有良好的重复性、准确度及专属性,可以用于p-ATG临床用药后血清浓度的检测。Objective To establish and verify the pharmacokineic detection method of porcine anil-human T cell immunoglobulin(p-ATG),providing technical service for the measurement of total serum p-ATG concentration in patients after clinical medication.Methods Using rabbit anti-pig IgG F(ab')2 as coating antibody,horseradish peroxidase-labeled goat anti-pig IgG as secondary antibody,a double-antibody sandwich ELISA for p-ATG detection in human serum was established.The standard curve fitting range,parallelism,precision,accuracy and specificity of the method were verified.Using the established method,clinical blood samples of 13 patients administered with different p-ATG doses were tested and the pharmacokinetics was analyzed.Results Starting at 0.44μg/ml initial concentration,21-27 times diluted standards concentrations and absorbance values showed an S curve with coefficient of determination(R2)>0.99 and recoveries in 89.91%-112.96%.The absolute value of coefficient of variation(CV)between parallelism sample slope and standard slope was<10%,R2>0.99 and CV of R2 was<5%.The inter-plate and intra-plate curves CVs were<10%.The repeatability CV was<10%and recoveries were 94.23%-101.80%.The spiked recoveries were 99.87%-104.94%.There were statistically significant differences between sera p-ATG concentrations of post-p-ATG-infused patients and those of healthy and pre-infused individuals(F=1.48,P<0.05).The clinical blood samples from 20,25 and 30 mg dosage groups showed that the p-ATG half-lives were 20.6,16.6 and 16.2 d,respectively.Conclusion The established method has good repeatability,accuracy and specificity,and can be used for the detection of serum concentration of p-ATG after clinical administration.

关 键 词：抗人T细胞猪免疫球蛋白 药代动力学 酶联免疫吸附测定 

分 类 号：R392[医药卫生—免疫学]