Hossain Faisal, Shen Qiming, Balasuriya Nicholas, Law John Lok Man, Logan Michael, Houghton Michael, Tyrrell D Lorne, Joyce Michael A, Serpe Michael J
Department of Chemistry, University of Alberta, Edmonton, Alberta T6G 2G2, Canada.
Department of Chemistry, Faculty of Science, University of Chittagong, Chattogram 4331, Bangladesh.
Anal Chem. 2023 May 16;95(19):7620-7629. doi: 10.1021/acs.analchem.3c00481. Epub 2023 May 7.
A sensor capable of quantifying both anti-SARS-CoV-2 spike receptor-binding domain (RBD) antibody levels and the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) virus in saliva and serum was developed. This was accomplished by exploiting the enzymatic reaction of maltose and orthophosphate (PO) in the presence of maltose phosphorylase to generate an equivalent amount of glucose that was detected using a commercial glucometer test strip and a potentiostat. Important for this approach is the ability to generate PO in an amount that is directly related to the concentration of the analytes. RBD-modified magnetic microparticles were used to capture anti-SARS-CoV-2 spike RBD antibodies, while particles modified with anti-SARS-CoV-2 nucleocapsid antibodies were used to capture SARS-CoV-2 nucleocapsid protein from inactivated virus samples. A magnet was used to isolate and purify the magnetic microparticles (with analyte attached), and alkaline phosphatase-conjugated secondary antibodies were bound to the analytes attached to the respective magnetic microparticles. Finally, through enzymatic reactions, specific amounts of PO (and subsequently glucose) were generated in proportion to the analyte concentration, which was then quantified using a commercial glucometer test strip. Utilizing glucose test strips makes the sensor relatively inexpensive, with a cost per test of ∼US $7 and ∼US $12 for quantifying anti-SARS-CoV-2 spike RBD antibody and SARS-CoV-2, respectively. Our sensor exhibited a limit of detection of 0.42 ng/mL for anti-SARS-CoV-2 spike RBD antibody, which is sensitive enough to quantify typical concentrations of antibodies in COVID-19-infected or vaccinated individuals (>1 μg/mL). The limit of detection for the SARS-CoV-2 virus is 300 pfu/mL (5.4 × 10 RNA copies/mL), which exceeds the performance recommended by the WHO (500 pfu/mL). In addition, the sensor exhibited good selectivity when challenged with competing analytes and could be used to quantify analytes in saliva and serum matrices with an accuracy of >94% compared to RT-qPCR.
开发了一种能够同时定量检测唾液和血清中抗SARS-CoV-2刺突受体结合域（RBD）抗体水平以及严重急性呼吸综合征冠状病毒2（SARS-CoV-2）病毒的传感器。这是通过在麦芽糖磷酸化酶存在的情况下利用麦芽糖和正磷酸盐（PO）的酶促反应来生成等量的葡萄糖实现的，该葡萄糖使用商用血糖仪测试条和恒电位仪进行检测。这种方法的关键在于能够生成与分析物浓度直接相关的PO量。RBD修饰的磁性微粒用于捕获抗SARS-CoV-2刺突RBD抗体，而用抗SARS-CoV-2核衣壳抗体修饰的微粒用于从灭活病毒样本中捕获SARS-CoV-2核衣壳蛋白。使用磁铁分离和纯化磁性微粒（附着有分析物），碱性磷酸酶偶联的二抗与附着在各自磁性微粒上的分析物结合。最后，通过酶促反应，根据分析物浓度生成特定量的PO（随后是葡萄糖），然后使用商用血糖仪测试条进行定量。使用葡萄糖测试条使该传感器相对便宜，定量抗SARS-CoV-2刺突RBD抗体和SARS-CoV-2的每次测试成本分别约为7美元和12美元。我们的传感器对抗SARS-CoV-2刺突RBD抗体的检测限为0.42 ng/mL，灵敏度足以定量COVID-19感染或接种疫苗个体中的典型抗体浓度（>1 μg/mL）。SARS-CoV-2病毒的检测限为300 pfu/mL（5.4×10 RNA拷贝/mL），超过了世界卫生组织推荐的性能（500 pfu/mL）。此外，该传感器在受到竞争性分析物挑战时表现出良好的选择性，与RT-qPCR相比，可用于定量唾液和血清基质中的分析物，准确度>94%。
