Cytotoxicity LDH Assay Kit-WST

Protocol tips

Upstream tips	Protocol tips	Downstream tips
		- If large variation of absorbance is present, please check for presence of bubble and break them; Mix reagents properly with accurate volumes as suggested. - Incase of High background values, use the serum-free medium or the medium contains the serum less than 5%.

Downstream tips
- If large variation of absorbance is present, please check for presence of bubble and break them; Mix reagents properly with accurate volumes as suggested. - Incase of High background values, use the serum-free medium or the medium contains the serum less than 5%.

Publication protocol

Cell death was evaluated colorimetrically by the activity of the lactate dehydrogenase (LDH) released in the media, using cytotoxicity LDH Assay Kit (Dojindo, Shanghai, China). Briefly, cells were treated with increasing concentrations of S17092 (0.026–130 μM) or vehicle (dimethylsulfoxide, DMSO) for 24 or 48 h, the liquid supernatant were collected and LDH activity was detected according to the manufacturer’s instruction; the absorbance (optical density, OD) at 490 nm was detected using a microplate reader (uQuant, Biotek, Winooski, VT). The average absorbance from each triplicate set of wells were calculated and subtract the background control value from each absorbance one and the percent cytotoxicity was expressed as a percentage of maximal reaction, which was obtained through the lysis of all cells. Cytotoxicity (%) = (sample OD − control OD) / (maximal reaction OD − control OD) ×100. The proliferation rate of the L02 cells was determined using a cell counting kit-8 (CCK-8, Dojindo, Shanghai, China) according to the manufacturer’s instructions. In brief, L02 cells were seeded on a 96-well plate at the density of 104 cells per well. Then, the cells were treated with increasing concentrations of S17092 (0.026–130 μM) or vehicle (dimethylsulfoxide, DMSO) for 24 or 48 h [15, 18]. CCK-8 was added, and the absorbance (optical density, OD) at 450 nm was detected using a microplate reader (uQuant, Biotek, Winooski, VT).

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