Infections and Cancer Epidemiology, Infection, Inflammation and Cancer Research Program, German Cancer Research Center (DKFZ), Heidelberg, Germany; BioNTech Corporation, Mainz, Germany.
Institut für Mikrobiologie und Hygiene, Universität des Saarlands, Homburg, Germany.
J Immunol Methods. 2019 Mar;466:47-51. doi: 10.1016/j.jim.2019.01.007. Epub 2019 Jan 18.
By inducing immunosuppression in infected patients, human immunodeficiency virus-1 (HIV-1) generates a favorable environment for opportunistic infections and the development of several human cancers. In order to detect individual serum or plasma HIV-1 antibody status for epidemiological studies, high-throughput HIV-1 Multiplex Serology was developed. Seven HIV-1 antigens were recombinantly expressed in E. coli as N-terminal glutathione-S-transferase (GST) fusion proteins that are bound to glutathione-coupled sets of beads with distinct fluorescent color. Combining all bead sets in a suspension array allowed for simultaneous detection of antibodies targeting structural, regulatory and accessory proteins expressed during HIV-1 infection. HIV-1 Multiplex Serology was validated with 244 reference sera whose HIV-1 serostatus had been pre-determined by screening microparticle immunoassay and confirmatory line immunoassay. The multifunctional protein GAG emerged as an excellent marker to determine HIV-1 serostatus with a specificity of 99% (95% CI 96%-100%) and sensitivity of 100% (95% CI 88%-100%). Seropositivity for multiple HIV-1 antigens appeared to be characteristic for HIV-1 infected individuals (median number of antigens recognized in reference assay positive sera: 4; median number of antigens recognized in reference assay negative sera: 0), indicating a broad immune response targeting also regulatory and accessory proteins which may be useful for the identification of antibody patterns specific for infection-associated disease stages. HIV-1 Multiplex Serology performs similarly to conventional HIV-1 serology but eliminates the need for a two-step screening approach with subsequent confirmation assay. Thus, this high-throughput method will facilitate large-scale epidemiological studies of the role of HIV-1 in cancer development.
通过诱导感染患者的免疫抑制，人类免疫缺陷病毒 1（HIV-1）为机会性感染和几种人类癌症的发展创造了有利环境。为了检测个体血清或血浆中 HIV-1 抗体的状态进行流行病学研究，开发了高通量 HIV-1 多重血清学检测。七种 HIV-1 抗原在大肠杆菌中作为 N 端谷胱甘肽-S-转移酶（GST）融合蛋白表达，与谷胱甘肽偶联的珠组结合，具有不同的荧光颜色。将所有珠组组合在悬浮阵列中，可同时检测针对 HIV-1 感染期间表达的结构、调节和辅助蛋白的抗体。使用 244 份参考血清对 HIV-1 多重血清学进行了验证，这些血清的 HIV-1 血清学状态已经通过微粒免疫测定和确认线免疫测定进行了预先确定。多功能蛋白 GAG 作为一种极好的标志物，其特异性为 99%（95%CI96%-100%），敏感性为 100%（95%CI88%-100%），可用于确定 HIV-1 血清学状态。针对多种 HIV-1 抗原的血清阳性似乎是 HIV-1 感染个体的特征（参考检测阳性血清中识别的抗原中位数：4；参考检测阴性血清中识别的抗原中位数：0），表明针对调节和辅助蛋白的广泛免疫反应也可能有助于识别与感染相关的疾病阶段特异性的抗体模式。HIV-1 多重血清学的性能与传统的 HIV-1 血清学相似，但消除了两步筛选方法的需要，随后是确认检测。因此，这种高通量方法将有助于大规模的 HIV-1 在癌症发展中的作用的流行病学研究。
