biostudies-literatureLiang ZGeneral Program of the National Natural Science Foundation of ChinaNational Key R&amp;D Program of ChinaBeijing Municipal Science and Technology ProjectChangping laboratorye2261566https://www.ebi.ac.uk/biostudies/studies/S-EPMC10540657biostudies-literatureUnknown12(2)<b>ABSTRACT</b>The global outbreak of COVID-19 has caused a severe threat to human health; therefore, simple, high-throughput neutralization assays are desirable for developing vaccines and drugs against COVID-19. In this study, a high-titre SARS-CoV-2 pseudovirus was successfully packaged by truncating the C-terminus of the SARS-CoV-2 spike protein by 21 amino acids and infecting 293 T cells that had been stably transfected with the angiotensin-converting enzyme 2 (ACE2) receptor and furin (named AF cells), to establish a simple, high-throughput, and automated 384-well plate neutralization assay. The method was optimized for cell amount, virus inoculation, incubation time, and detection time. The automated assay showed good sensitivity, accuracy, reproducibility, Z' factor, and a good corEmerging microbes & infectionsDevelopment of an automated, high-throughput SARS-CoV-2 neutralization assay based on a pseudotyped virus using a vesicular stomatitis virus (VSV) vector.PMC1054065782172244 &amp; 32070678Z2111000025210182021YFC08633002021YFC23025002021YFC2301700Wu JZhao CLiang ZNie JLiu SZhang LLi THuang WTong JQin HWang YLu QWu XYu YfalseDevelopment of an automated, high-throughput SARS-CoV-2 neutralization assay based on a pseudotyped virus using a vesicular stomatitis virus (VSV) vector.<b>ABSTRACT</b>The global outbreak of COVID-19 has caused a severe threat to human health; therefore, simple, high-throughput neutralization assays are desirable for developing vaccines and drugs against COVID-19. In this study, a high-titre SARS-CoV-2 pseudovirus was successfully packaged by truncating the C-terminus of the SARS-CoV-2 spike protein by 21 amino acids and infecting 293 T cells that had been stably transfected with the angiotensin-converting enzyme 2 (ACE2) receptor and furin (named AF cells), to establish a simple, high-throughput, and automated 384-well plate neutralization assay. The method was optimized for cell amount, virus inoculation, incubation time, and detection time. The automated assay showed good sensitivity, accuracy, reproducibility, Z' factor, and a good cor2023-01-01T00:00:00Z2023 Dec2026-05-29T01:58:01.725Z2025-02-19T00:01:06.723ZS-EPMC105406573772710710.1080/22221751.2023.2261566