人成熟型转化生长因子-β_1融合蛋白的纯化及其多克隆抗体的制备与初步临床应用  

Purification of human TGF-β_1 fusion protein and clinical detection of TGF-β_1 by its polyclonal antibody

作　　者：龚环宇[1] 胡国龄[2] 谭德明[2] 侯珏[3] 

机构地区：[1]中南大学湘雅三医院,湖南长沙410013 [2]中南大学湘雅医院,湖南长沙410008 [3]中南大学湘雅医学院,湖南长沙410013

出　　处：《中国感染控制杂志》2007年第6期372-375,400,共5页Chinese Journal of Infection Control

摘　　要：目的获得兔抗人成熟型转化生长因子-β1(TGF-β1)多克隆抗体,并将其初步应用于临床检测。方法将人成熟型TGF-β1融合蛋白经亲和层析法纯化,并将其作为抗原免疫家兔,获得兔抗血清,经饱和硫酸铵纯化,获得初步纯化的兔抗人TGF-β1多克隆抗体。采用双抗夹心间接酶联免疫吸附试验(ELISA)测定健康对照者、慢性肝病及肝硬化患者血清TGF-β1的OD值。结果获得初步纯化的兔抗人TGF-β1特异性多克隆抗体,该抗体能与人TGF-β1起反应。各组血清TGF-β1的检测OD450值为:轻、中度慢性肝炎组11.333±6.136,慢性重症肝炎组16.400±6.959,肝硬化组14.975±6.090,健康对照组7.275±3.222;前三组分别与健康对照组比较,差异均有显著性(P<0.01);慢性重症肝炎组、肝硬化组分别与轻、中度慢性肝炎组比较,差异亦有显著性(P<0.01)。结论制备的TGF-β1多克隆抗体可初步用于临床检测,判断肝纤维化程度。Objective To obtain the fusion protein of transforming growth factor beta 1 （TGF-β1） and the rabbit anti-human TGF-β1 polyclonal antibody, and to detect the serum in patients by this polyclonal antibody. Methods The fusion protein was purified by affinity chromatography, two rabbits were immunized with the purified fusion protein, and the immune sera of rabbits were collected. Antibodies obtained from the immune sera were purified by saturated ammonium sulfate, and the indirect sandwich enzyme immunoassay was used to detect the OD values of TGF-β1 in sera of healthy control and patients with chronic liver diseases and cirrhosis. Results The purified TGF- β1 fusion protein was obtained, the purified specific polyclonal antibody of rabbit-anti-human TGF-β1 was also obtained from the immune sera of the rabbits, and could respond to human TGF-β1. The values of OD451, in every group were as follows： 11. 333 ± 6. 136 in slight and moderate chronic hepatitis groups, 16. 400 ± 6. 959 in chronic severe hepatitis group, 14. 975 ± 6. 090 in hepatocirrhosis group,7. 275 ± 3. 222 in healthy control group, there was significant difference in OD450, values between the former three groups and healthy group respectively （P〈0. 01）; there were also obvious difference between chronic severe hepatitis, hepatocirrhosis groups and slight, moderate chronic hepatitis groups respectively（P〈0. 01）. Conclusion The rabbit-anti-human TGF-β1 polyclonal antibody can be used in the diagnosis of initial stage of hepatic fibrosis.

关 键 词：人成熟型转化生长因子-β1 融合蛋白 亲和层析法 多克隆抗体 肝纤维化 实验室技术与方法 

分 类 号：R446.61[医药卫生—诊断学]