| Assay Method Information | |
| | Human Cathepsin L (CatL) Enzymatic Assay |
| Description: | Human cathepsin L enzymatic assay was performed in assay buffer (50 mM MES pH 5.5, 2.5 mM DTT, 0.5 mM EDTA) to assess the inhibition of human CatL by tested compounds. 60 μL of the compounds were added to 384-well dilution plate. The compound solution was diluted for 1:3 in succession in DMSO for each column for 10 pts. 0.05 μL diluted compound solution was added in each row to 384 assay plates (Corning 4514) using Echo (LABCYTE 655), each column containing 2 replicates. 5 μL working solution of human CatL enzyme (Abcam #ab81780) was added to 384-well assay plate, centrifuge 1000 rpm for 1 min. The mixture was incubated at 2500 for 15 min, then add 5 μL working solution of CatL substrate (Genscript #C7360HB140_5) to initiate the reaction (CatL: 0.05 nM, Substrate: 500 nM). Incubate at 2500 for 30 min. Reading Ex370 nm and Em460 nm fluorescence signals with BMG CLARIO Star Plusaucu. Percent inhibition for each compound was calculated and IC50 (half maximal inhibitory concentration) was fitted from non-linear regression by XLfit 5.5.0. |
| Affinity data for this assay | |
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