Blocking antibodies against SARS-CoV-2 RBD isolated from a phage display antibody library using a competitive biopanning strategy

Xin Zeng; Lingfang Li; Jing Lin; Xinlei Li; Bin Liu; Yang Kong; Shunze Zeng; Jianhua Du; Huahong Xiao; Tao Zhang; Shelin Zhang; Jianghai Liu

The neutralization effects of antibodies on SARS-CoV-2 pseudovirus were performed by the Genscript Inc. (Nanjing, China) under a research service contract. Briefly, 20,000 of the human ACE2-overexpressing Hela monoclonal cells were seeded into each well of a 96-well plate. SARS-CoV-2 pseudovirus and antibodies were incubated at ambient temperature for 1 hour. The mixture was transferred into wells and incubated with cells at 37°C, 5% CO2 for 24 hours. The culture medium was freshly replaced, and cells were incubated for another 24 hours. The culture medium was removed, and cells were rinsed with PBS. 50µl lysis buffer was added and further incubated at ambient temperature for 40 minutes. 40µl supernatant was transferred to a Sterile Un-Clear 96-well plate with the Bio-Glo luciferase substrate added, and the luminescence signal was measured with EnVision. The dose response curves were plotted with the relative luminescence unit against the antibody concentration. The assay results were processed by Microsoft Office Excel 2013 and GraphPad Prism 6.