SARS-CoV-2 specific antibody responses in COVID-19 patients

NISREEN M.A. OKBA; Marcel A Muller; Wentao Li; Chunyan Wang; Corine H. GeurtsvanKessel; Victor M. Corman; Mart M. Lamers; Reina S. Sikkema; Erwin de Bruin; Felicity D. Chandler; Yazdan Yazdanpanah; Quentin Le Hingrat; Diane Descamps; Nadhira Houhou-Fidouh; Chantal B. E. M. Reusken; Berend-Jan Bosch; Christian Drosten; Marion P.G. Koopmans; Bart L. Haagmans

PRNT was used as a reference for this study, because neutralization assays are the standard for CoV serology. We tested serum samples for their neutralization capacity against SARS-CoV-2 (German isolate; GISAID ID EPI_ISL 406862; European Virus Archive Global # 026V-03883) by plaque-reduction neutralization test (PRNT) as previously described for with some modifications (9) . Heat-inactivated samples were 2-fold serially diluted in DMEM medium supplemented with NaHCO3, HEPES buffer, penicillin, streptomycin, and 1% fetal bovine serum, starting at a dilution of 1:10 in 50 μL. Fifty μL of the virus suspension (400 spot forming units) was added to each well and incubated at 37°C for 1 h. Following incubation, the mixtures were added on Vero-E6 cells and incubated at 37°C for 1 more hour. The cells were then washed and further incubated in medium for 8 h. After the incubation, the cells were fixed and stained with polyclonal rabbit anti-SARS-CoV antibody (Sino Biological). The cells were then fixed and stained using a rabbit anti-SARS-CoV serum and a secondary peroxidase-labelled goat antirabbit IgG (Dako). The signal was developed using a precipitate forming TMB substrate (True Blue, KPL) and the number of infected cells per well were counted using the ImmunoSpot® All rights reserved. No reuse allowed without permission.