| Assay Method Information | |
| | Inhibition of 3CL Protease (3CLPro) of SARS-CoV-2 |
| Description: | To study the inhibitory activities against SARS-CoV-2 3CLPro of test compounds, an assay was determined in vitro by measuring the enhanced fluorescence due to cleavage of the fluorogenic substrate (Dabcyl-KTSAVLQSGFRKME-Edans). For analyzing the inhibition potential, various compounds were dissolved in 1, 8 or 40% dimethyl sulfoxide (DMSO) aqueous solution as the compound stocks. Different concentration of each stocks (5 μl) was pre-incubated with 45 μl reaction mixture (50 nM SARS-CoV-2 viral 3CL protease in 20 mM Bis-Tris, pH 7.4) at 37° C. for 30 minutes. Afterwards, 50 μl of the fluorogenic peptide substrate (6 μM) was added into the mixture and gently mixed to get the final DMSO concentration 0.05, 0.40 or 2.00% solution. The difference of fluorescence intensity resulting from the reaction was measured at 485 nm with excitation at 360 nm using a fluorescence plate reader at 37° C. for 4 min. The protease activity was presented as fluorescence intensity and calculated by the following equation:Inhibition (%)=1−[(fluorescencesample, 4 min−fluorescencesample, 0 min)/(fluorescenceddH2O, 4 min−fluorescenceddH2O, 0 min)]×100%. |
| Affinity data for this assay | |
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