摘要
嗜酸性粒细胞阳离子蛋白(ECP)是一组由不同种类组成的碱性蛋白质,被广泛应用于过敏性患者临床检验的标志分子,也是酸性粒细胞活性和更新的一个指标。
本实验以ECP为抗原,制备了多克隆抗体和单克隆抗体,经辛酸-硫酸铵和蛋白亲和层析及浓缩后,多克隆抗体和单克隆抗体效价均达到1:10~7。初步建立了基于ECP的双抗夹心ELISA方法,其检测范围为10ng/ml-900ng/ml,检测下限为5ng/ml。
将方法应用于戊型肝炎患者血清样本进行检测,其中:戊型抗原Ag阳性样本9个,IgG阳性样本100个,IgM阳性样本72个,普通健康人群血清样本10个。检测结果:戊肝阴性样本ECP水平最高,其次为戊肝IgG阳性样本,Ag阳性样本ECP水平较低,IgM阳性样品ECP水平最低。t检验结果显示:IgM、Ag阳性样本两者差异显著,有统计学意义(P＜0.005),而戊肝IgG阳性样本与戊肝阴性样本无统计学意义(0.10＜P＜0.25),戊型肝炎病毒可能抑制了嗜酸性粒细胞的活性,因而戊型肝炎早期阳性血清样本中ECP的含量较低,感染后期接近健康水平阳性血清样本中ECP的含量接近阴性血清,该实验结果国内首次报道。
本结果为进一步研究嗜酸性粒细胞ECP的释放水平与戊型肝炎病毒感染的相关性与及做为戊型肝炎的一种早期诊断标志物提供了实验基础。
Eosinophile cationic protein(ECP) is composed of a variety of Basic Proteins,which has been widely used as the symbol molecule for clinical examination of allergic patients and also the index for eosinophile activity and improvement.
Taking ECP as antigen,the experiment has made polyclonal antibody and monoclonal antibody;after affinity chromatography and condensation ware performed with Caprylic acid -ammonium sulphate and protein,the valence of polyclonal antibody and monoclonal antibody reachedl:10~(7.) Based on ECP,the High efficient and specific ELISA method was preliminarily established.The testing scope is 10ng/ml-900ng/ml;lower limit of testing is 5ng/ml.
The blood serum samples of Hepatitis E patient ware tested by this method including 9 Ag positive samples,100 IgG positive samples,72 IgM positive samples,10 serum samples of healthy ordinary persons.The testing data shows:Hepatitis E negative sample contain the highest ECP,the second is Hepatitis E IgG positive sample;Ag positive sample contain lower ECP;IgM positive sample contain the lowest ECP;the testing result shows:there is obvious variance between IgM positive sample and Ag positive sample,which is of statistics meaning(P＜0.005) while IgG positive sample and Hepatitis E negative sample has no statistics meaning(0.10＜P＜0.25 ).The result indicates: Hepatitis E virus may restrain the activity of Eosinophile,thus the prophase positive blood serum sample of Hepatitis E contain low ECP and the positive blood serum sample close to healthy condition in later phase contain higher ECP.The testing result has been the firstly reported in China.
The testing result provides experimental basis for further study on the correlation between the ECP release level and Hepatitis E virus infection as well as the prophase diagnosis symbol for Hepatitis E.
引文
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