| Assay Method Information | |
| | Enzyme Activity Assay |
| Description: | An inhibition test was conducted using 200 nM recombinant SARS-CoV-2 main protease and 15 μM fluorescent substrate (Dabcyl-TSAVL QSGFRK-Glu (EDANS); Genscript). An assay buffer consisted of 50 mM Tris-HCl and 1 mM EDTA at pH 7.3. SARS-CoV-2 3CLpro was dissolved in 25 μL of the assay buffer and mixed with compounds at different concentrations. The mixture was incubated at 37° C. for 30 minutes. A substrate dissolved in 25 μL of the assay buffer was then added to initiate a reaction. Fluorescent signals at 350 nm (excitation)/490 nm (emission) were measured immediately each minute in 10 minutes at 37° C. using a SpectraMax® M5 multimode microplate reader. RFU at 6th minute of the reaction with the compounds at different concentrations in contrast with a reaction with the lowest concentration was used to generate an IC50 curve. IC50 values corresponding to SARS-CoV-2 3CLpro were measured for each compound at 12 concentrations. |
| Affinity data for this assay | |
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