*2.3. Cell Viability Assay and E*ffi*cacy Study of dec-RVKR-cmk*

The cytotoxic concentration 50 (CC50) of dec-RVKR-cmk was determined using the CellTiter-GLO One Solution Assay kit (Promega). This assay was used to detect the viability of cultured cells on the basis of ATP quantification of cells. Briefly, Vero and C6/36 cells were seeded (10,000 cells per well) in a 96-well plate, 24 h before compound treatment. Culture supernatants were replaced with different concentrations of dec-RVKR-cmk or DMSO. Each concentration was tested in triplicate. After 72 h, cells were washed with phosphate-buffered saline (PBS) and an equal volume of (100 μL) CellTiter-GLO reagent was added to each well. For appropriate cell lysis, cells were agitated in a shaker for 2 min and then incubated for 10 min at room temperature. A multimode plate reader was used to quantitate luminescence signals in each condition and then the luminescence value was compared with its corresponding DMSO control. The efficacy of dec-RVKR-cmk against ZIKV (0.2 MOI) and JEV (0.2 MOI) was studied by using different concentrations (1, 10, 50, and 100 μM). The inhibitory concentration 50 (IC50) of dec-RVKR-cmk was determined by counting visible plaques produced by ZIKV or JEV. Both CC50 and IC50 were calculated by non-linear regression model using GraphPad prism7.
