Detection of HIV-1 p24 antigen using streptavidin-biotin and gold nanoparticles based immunoassay by inductively coupled plasma mass spectrometry
Detection of HIV-1 p24 antigen using streptavidin-biotin and gold nanoparticles based immunoassay by inductively coupled plasma mass spectrometry
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使用基于链霉亲和素-生物素和金纳米颗粒的电感耦合等离子体质谱免疫测定法检测 HIV-1 p24 抗原
DOI：
10.1039/c4ja00026a
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发表时间：
2014-08-01
影响因子：
3.4
通讯作者：
Hu, Shenghong
中科院分区：
文献类型：
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作者：
He, Qian;Zhu, Zhenli;Hu, Shenghong
采用生物素-链亲和素（BA）体系和金纳米粒子（Au NPs）免疫分析法，建立了一种电感耦合等离子体质谱（ICP-MS）检测HIV-1 p24抗原的灵敏方法。在这种免疫分析中，p24抗原首先被固定的抗hiv -1 p24单克隆抗体捕获。与生物素化抗p24多克隆抗体和Au NPs标记的链亲和素免疫反应后，用稀释的HNO3 （5%, v/v）解离Au NPs，然后将其引入ICP-MS进行测量。在优化条件下，p24抗原在7.5 ~ 75 pg mL（-1）范围内呈线性关系，检出限为1.49 pg mL(-1) （3 sigma, n = 5）。p24抗原37.5 pg mL(-1) 3次重复测量的相对标准偏差（RSD）为3.7%。其他蛋白如人IgG、人HSA、人CEA和人AFP对p24抗原检测无明显干扰。该方法也适用于人工阳性人血清样品中p24浓度的测定。与生物素-链亲和素酶联免疫吸附法（BA-ELISA）检测p24抗原相比，ICP-MS联免疫分析方法处理的是Au NPs标记的抗体，而不是酶偶联抗体，因此不使用有毒的酶底物试剂。另外，省去了显色步骤，简化了实验过程，节省了实验时间。该方法为检测HIV-1 p24抗原提供了一种灵敏的方法。
A sensitive assay for detection of HIV-1 p24 antigen by inductively coupled plasma mass spectrometry (ICP-MS) was developed using a biotin-streptavidin (BA) system and gold nanoparticles (Au NPs) based immunoassay. In this immunoassay, the p24 antigen was firstly captured by an immobilized anti-HIV-1 p24 monoclonal antibody. After immunoreactions with the biotinylated anti-p24 polyclonal antibody and Au NPs-labeled streptavidin, diluted HNO3 (5%, v/v) was used to dissociate Au NPs, which were then introduced to the ICP-MS for measurements. Under the optimized conditions, the calibration graph for the p24 antigen was linear in the range of 7.5-75 pg mL(-1) with a detection limit of 1.49 pg mL(-1) (3 sigma, n = 5). The relative standard deviation (RSD) for three replicate measurements of 37.5 pg mL(-1) of the p24 antigen was 3.7%. Other proteins, such as human IgG, human HSA, human CEA and human AFP, did not obviously interfere with the assay for p24 antigen. This method was also applied to measure p24 concentrations in artificially positive human serum samples. Compared with the biotin-streptavidin enzyme-linked immunosorbent assay (BA-ELISA) method for p24 antigen detection, the ICP-MS linked immunoassay process deals with Au NPs - tagged instead of enzyme-conjugated antibodies, making it free of toxic enzyme substrate reagents. In addition, it also simplifies the experimental process and saves the experimental time, since the color rendering steps are omitted. The proposed approach provides a sensitive method for HIV-1 p24 antigen determination.