A simple and high-throughput luciferase immunosorbent assay for both qualitative and semi-quantitative detection of anti-HIV-1 antibodies
A simple and high-throughput luciferase immunosorbent assay for both qualitative and semi-quantitative detection of anti-HIV-1 antibodies
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一种简单且高通量的荧光素酶免疫吸附测定，用于定性和半定量检测抗 HIV-1 抗体
DOI：
10.1016/j.virusres.2018.12.017
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发表时间：
2019-04-02
期刊：
影响因子：
5
通讯作者：
Tang, Shixing
中科院分区：
文献类型：
--
作者：
Wang, Haiying;Cai, Qundi;Tang, Shixing
在这项研究中，我们描述了一种高灵敏度和高通量的荧光素酶免疫吸附试验(LISA)，用于定性和定量检测抗HIV-1抗体。用蛋白A/G包被的微孔板捕获血清或血浆中的抗HIV抗体，并用表达纳米荧光素酶(Nluc)酶的粗细胞裂解物与HIV-1p24或gp41抗原融合，不需要蛋白纯化。加入呋喃西林底物后，以荧光素酶发光单位定量检测抗HIV抗体。LISA的检测线性范围很宽，其灵敏度约为ELISA的10(4)倍。LISA检测抗p24和抗gp41的敏感性分别为99.5%和100%，特异度相同(100%)。LISA还可以监测接受抗逆转录病毒治疗(ART)的个体随着时间的推移抗HIV-1抗体反应的变化，并能够充分区分最近和长期的HIV-1感染。我们的初步结果表明，LISA可能为同时检测HIV-1、定量检测抗HIV抗体以及区分HIV-1感染阶段提供一个新的通用免疫分析平台。
In this study, we described an ultrasensitive and high-throughput luciferase immunosorbent assay (LISA) for qualitative and quantitative detection of anti-HIV-1 antibody. Anti-HIV antibody in serum or plasma samples was captured by protein A/G-coated microtiter plate and detected with crude cell lysates expressing Nanoluc luciferase (Nluc) enzyme fused with HIV-1 p24 or gp41 antigen without the need of protein purification. After the addition of furimazine substrate, anti-HIV antibodies were quantitatively measured as luciferase light units. LISA showed a wide linear range of detection and was about 10(4)-fold more sensitive than ELISA. For the detection of both anti-p24 and anti-gp41, LISA showed extraordinary sensitivity (99.5% and 100%, respectively) and equivalent specificity (100%). LISA could also monitor the change in the anti-HIV-1 antibody response over time in antiretroviral therapy (ART) treated individuals, and can sufficiently distinguish between recent and long-term HIV-1 infections. Our preliminary results indicate that LISA may provide a novel universal immunoassay platform for simultaneous HIV-1 detection, quantitative measurement of anti-HIV antibodies as well as the differentiation of HIV-1 infection stages.