National Center of Infectious and Parasitic Diseases, 44A "Gen. Stoletov" Blvd., 1233 Sofia, Bulgaria.
Institute of Optical Materials and Technologies "Acad. J. Malinowski" (IOMT), Bulgarian Academy of Sciences (BAS), 109 "Acad. G. Bonchev" Str., 1113 Sofia, Bulgaria.
Biosensors (Basel). 2021 Jul 26;11(8):251. doi: 10.3390/bios11080251.
Cell-based assays are a valuable tool for examination of virus-host cell interactions and drug discovery processes, allowing for a more physiological setting compared to biochemical assays. Despite the fact that cell-based SPR assays are label-free and thus provide all the associated benefits, they have never been used to study viral growth kinetics and to predict drug antiviral response in cells. In this study, we prove the concept that the cell-based SPR assay can be applied in the kinetic analysis of the early stages of viral infection of cells and the antiviral drug activity in the infected cells. For this purpose, cells immobilized on the SPR slides were infected with human coronavirus HCov-229E and treated with hydroxychloroquine. The SPR response was measured at different time intervals within the early stages of infection. Methyl Thiazolyl Tetrazolium (MTT) assay was used to provide the reference data. We found that the results of the SPR and MTT assays were consistent, and SPR is a reliable tool in investigating virus-host cell interaction and the mechanism of action of viral inhibitors. SPR assay was more sensitive and accurate in the first hours of infection within the first replication cycle, whereas the MTT assay was not so effective. After the second replication cycle, noise was generated by the destruction of the cell layer and by the remnants of dead cells, and masks useful SPR signals.
基于细胞的测定法是研究病毒-宿主细胞相互作用和药物发现过程的有价值的工具，与生化测定法相比，它提供了更接近生理的环境。尽管基于细胞的 SPR 测定法是无标记的，因此提供了所有相关的好处，但它们从未被用于研究病毒的生长动力学和预测细胞中的药物抗病毒反应。在这项研究中，我们证明了这样一个概念，即基于细胞的 SPR 测定法可应用于细胞感染病毒的早期阶段的动力学分析以及感染细胞中的抗病毒药物活性。为此，将固定在 SPR 载玻片上的细胞用人类冠状病毒 HCov-229E 感染并用羟氯喹处理。在感染的早期阶段，以不同的时间间隔测量 SPR 响应。使用甲基噻唑基四唑 (MTT) 测定法提供参考数据。我们发现 SPR 和 MTT 测定法的结果是一致的，并且 SPR 是研究病毒-宿主细胞相互作用和病毒抑制剂作用机制的可靠工具。在第一个复制周期内感染的最初几个小时内，SPR 测定法比 MTT 测定法更敏感和准确，而 MTT 测定法则不那么有效。在第二个复制周期之后，细胞层的破坏和死细胞的残留物会产生噪声，从而掩盖了有用的 SPR 信号。
