基于化学发光动力学分辨策略的多组分免疫分析方法检测免疫球蛋白
发布时间：2019-03-24 15:38
【摘要】：化学发光免疫分析法(Chemiluminescence immunoassay, CLIA)是将化学发光反应和免疫反应相互结合,具有高灵敏度、高特异性,以放射免疫分析为理论基础建立起来的,用来检测抗原或者抗体的一种非放射标记免疫分析法。这种方法具有无需光源、仪器设备简单、灵敏度高、特异性强、线性范围宽、无放射性污染、可实现自动化等优点。多组分免疫分析(Multiplexed immunoassay, MIA)是目前临床诊断、环境监测、生物分析等领域的研究热点,具有分析成本低、分析通量高、所需时间短、样品消耗少等突出优点。近年来,大多数报道的MIA方法通常是基于空间分辨或多标记模式。本文基于CLIA,结合多标记模式构建了检测两种免疫球蛋白的新方法,具体开展了以下工作：本文以新的化学发光反应动力学分辨策略为基础,提出了一种近同时检测两种免疫球蛋白的多组分免疫分析方法。该方法利用化学发光动力学特征具有明显差异的两个化学发光标记物—吖啶酯和碱性磷酸酶作为信号探针,分别标记羊抗鼠IgG和兔抗鼠IgM,形成两个免疫复合物,通过竞争免疫分析模式,对两个分析物小鼠IgG (Mouse IgG, MIgG)和小鼠IgM(Mouse IgM, MIgM)进行检测。当加入共反应剂时,具有闪光型和辉光型动力学特征的两个反应同时被触发,在反应后0.2 s和500 s时,分别记录这两个反应的化学发光信号。该多组分免疫分析方法对于MIgG和MIgM的检测范围均为0.50-200 ng mL-1,检测限为0.16 ngmL-1 (S/N=3)。所构建的方法没有明显的信号重叠,并能够成功应用于检测小鼠血清样品中MIgG和MIgM,检测结果与商业化的ELISA方法检测结果相一致。该方法具有灵敏度、特异性较高和操作简便等优势,在一些如药物筛选,食品安全,环境监测和临床诊断等领域有很好的应用潜力。
[Abstract]:Chemiluminescent immunoassay (Chemiluminescence immunoassay, CLIA), which combines chemiluminescence reaction with immune reaction, has high sensitivity and specificity, and is based on the theory of RIA. A non-radiolabeled immunoassay used to detect antigens or antibodies. This method has the advantages of no light source, simple instrument and equipment, high sensitivity, strong specificity, wide linear range, no radioactive pollution and automation. Multi-component immunoassay (Multiplexed immunoassay, MIA) is a hot spot in the fields of clinical diagnosis, environmental monitoring, biological analysis and so on. It has the advantages of low analysis cost, high analysis flux, short time and low sample consumption. In recent years, most reported MIA methods are based on spatial resolution or multi-marker patterns. In this paper, a new method for detecting two kinds of immunoglobulin based on CLIA, combined with multi-labeled pattern is proposed. The following works are carried out in detail: based on the new chemiluminescence kinetic resolution strategy, A multi-component immunoassay method for near-simultaneous detection of two kinds of immunoglobulin was proposed. Using acridine ester and alkaline phosphatase as signal probes, two chemiluminescence markers, acridine ester and alkaline phosphatase, were used to label goat anti-mouse IgG and rabbit anti-mouse IgM, to form two immune complexes. Two analytes, mouse IgG (Mouse IgG, MIgG) and mouse IgM (Mouse IgM, MIgM), were detected by competitive immunoassays. When the co-reactant was added, the two reactions with the kinetic characteristics of flash and glow type were triggered at the same time. The chemiluminescence signals of the two reactions were recorded at 0.2 s and 500 s after the reaction, respectively. The detection range of both MIgG and MIgM is 0.50,200 ngmL-1 and the detection limit is 0.16 ngmL-1 (S/N=3). The constructed method has no obvious signal overlap and can be successfully applied to the detection of MIgG and MIgM, in mouse serum samples. The results are consistent with those of the commercial ELISA method. The method has the advantages of sensitivity, high specificity and simple operation. It has good application potential in some fields such as drug screening, food safety, environmental monitoring and clinical diagnosis.
【学位授予单位】：西南大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：R917
本文编号：2446455
[Abstract]:Chemiluminescent immunoassay (Chemiluminescence immunoassay, CLIA), which combines chemiluminescence reaction with immune reaction, has high sensitivity and specificity, and is based on the theory of RIA. A non-radiolabeled immunoassay used to detect antigens or antibodies. This method has the advantages of no light source, simple instrument and equipment, high sensitivity, strong specificity, wide linear range, no radioactive pollution and automation. Multi-component immunoassay (Multiplexed immunoassay, MIA) is a hot spot in the fields of clinical diagnosis, environmental monitoring, biological analysis and so on. It has the advantages of low analysis cost, high analysis flux, short time and low sample consumption. In recent years, most reported MIA methods are based on spatial resolution or multi-marker patterns. In this paper, a new method for detecting two kinds of immunoglobulin based on CLIA, combined with multi-labeled pattern is proposed. The following works are carried out in detail: based on the new chemiluminescence kinetic resolution strategy, A multi-component immunoassay method for near-simultaneous detection of two kinds of immunoglobulin was proposed. Using acridine ester and alkaline phosphatase as signal probes, two chemiluminescence markers, acridine ester and alkaline phosphatase, were used to label goat anti-mouse IgG and rabbit anti-mouse IgM, to form two immune complexes. Two analytes, mouse IgG (Mouse IgG, MIgG) and mouse IgM (Mouse IgM, MIgM), were detected by competitive immunoassays. When the co-reactant was added, the two reactions with the kinetic characteristics of flash and glow type were triggered at the same time. The chemiluminescence signals of the two reactions were recorded at 0.2 s and 500 s after the reaction, respectively. The detection range of both MIgG and MIgM is 0.50,200 ngmL-1 and the detection limit is 0.16 ngmL-1 (S/N=3). The constructed method has no obvious signal overlap and can be successfully applied to the detection of MIgG and MIgM, in mouse serum samples. The results are consistent with those of the commercial ELISA method. The method has the advantages of sensitivity, high specificity and simple operation. It has good application potential in some fields such as drug screening, food safety, environmental monitoring and clinical diagnosis.
【学位授予单位】：西南大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：R917
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