嗜酸性粒细胞阳离子蛋白抗体的制备及初步应用
发布时间：2018-10-11 07:45
【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。
[Abstract]:Eosinophil cationic protein (ECP) is a group of basic proteins composed of different species. It is widely used as a marker for clinical examination in allergic patients and is also an indicator of activity and renewal of acidophilic granulocytes. In this study, polyclonal antibodies and monoclonal antibodies were prepared using ECP as antigen. The titers of polyclonal antibodies and monoclonal antibodies reached 1: 10107after affinity chromatography and concentration of caprylic acid-ammonium sulfate and protein. A double antibody sandwich ELISA method based on ECP is established. The detection range is 10 ng / ml-900ng / ml and the detection limit is 5 ng / ml. The method was applied to the detection of hepatitis E patients' serum samples, including 9 positive samples of Ag, 100 positive samples of IgG, 72 positive samples of IgM, and 10 samples of serum from healthy people. The results showed that ECP level was the highest in hepatitis E negative samples, followed by IgG positive samples in hepatitis E, ECP level in Ag positive samples was lower, ECP level in IgM positive samples was the lowest. T test results showed that IgM,Ag positive samples had significant difference between them. There was statistical significance (P < 0.005), but there was no significant difference between IgG positive samples and negative samples (0.10 < P < 0.005). Hepatitis E virus might inhibit the activity of eosinophils. As a result, the content of ECP in the early positive serum samples of hepatitis E was lower than that in the positive serum samples at the later stage of infection, and the content of ECP in the positive serum samples was close to the negative serum. The results of the experiment were reported for the first time in China. The results provide experimental basis for further study on the correlation between eosinophil ECP release level and hepatitis E virus infection and as an early diagnostic marker for hepatitis E.
【学位授予单位】：长春理工大学
【学位级别】：硕士
【学位授予年份】：2010
【分类号】：R392
本文编号：2263403
[Abstract]:Eosinophil cationic protein (ECP) is a group of basic proteins composed of different species. It is widely used as a marker for clinical examination in allergic patients and is also an indicator of activity and renewal of acidophilic granulocytes. In this study, polyclonal antibodies and monoclonal antibodies were prepared using ECP as antigen. The titers of polyclonal antibodies and monoclonal antibodies reached 1: 10107after affinity chromatography and concentration of caprylic acid-ammonium sulfate and protein. A double antibody sandwich ELISA method based on ECP is established. The detection range is 10 ng / ml-900ng / ml and the detection limit is 5 ng / ml. The method was applied to the detection of hepatitis E patients' serum samples, including 9 positive samples of Ag, 100 positive samples of IgG, 72 positive samples of IgM, and 10 samples of serum from healthy people. The results showed that ECP level was the highest in hepatitis E negative samples, followed by IgG positive samples in hepatitis E, ECP level in Ag positive samples was lower, ECP level in IgM positive samples was the lowest. T test results showed that IgM,Ag positive samples had significant difference between them. There was statistical significance (P < 0.005), but there was no significant difference between IgG positive samples and negative samples (0.10 < P < 0.005). Hepatitis E virus might inhibit the activity of eosinophils. As a result, the content of ECP in the early positive serum samples of hepatitis E was lower than that in the positive serum samples at the later stage of infection, and the content of ECP in the positive serum samples was close to the negative serum. The results of the experiment were reported for the first time in China. The results provide experimental basis for further study on the correlation between eosinophil ECP release level and hepatitis E virus infection and as an early diagnostic marker for hepatitis E.
【学位授予单位】：长春理工大学
【学位级别】：硕士
【学位授予年份】：2010
【分类号】：R392
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