Assay Method Information

Assay Name:  In vitro antiviral cell-based assay (live SARS-CoV-2 IFA)
Description:  SARS-CoV-2 was provided by Korea Centers for Disease Control and Prevention (KCDC) . Vero cells were acquired from the ATCC and maintained in the Dulbecco s Modified Eagle s Medium (DMEM) supplemented with 10%FBS, and 1%Antibiotic-Antimycotic solution. DMEM supplemented with 2%FBS and 1%Antibiotic-Antimycotic solution are used as the assay medium. The main reagents used in this assay are Anti-SARS-CoV-2 N protein antibody, Alexa Fluor 488 goat anti-rabbit IgG (H + L) secondary antibody and Hoechst 33342. Ten-point dose-response curves (DRC) are generated for each compound. Vero cells are seeded at 1.2 10 4 cells per well in black 384-well, μClear plates (Greiner Bio-One) , 24 h prior to the experiment. For viral infection, SARS-CoV-2 is added at a multiplicity of infection (MOI) of ~0.0125. The cells are fixed at 24 hpi with 4%paraformaldehyde and analyzed by immunofluorescence. The acquired images are analyzed using software to quantify cell numbers and infection ratios, and antiviral activity is normalized to positive (mock) and negative (0.5%DMSO) controls in each assay plate. DRCs are fitted by sigmoidal dose-response models using XLfit 4 Software or Prism.
Affinity data for this assay
 

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