LIVE/DEAD™ Viability/Cytotoxicity Kit, for mammalian cells

Protocol tips

Upstream tips	Protocol tips	Downstream tips
Wash cells with PBS for 2-3 times	Add calcein AM: 2 μM; EthD-1: 4 μM for 40 minutes at room temperature

Upstream tips
Wash cells with PBS for 2-3 times
Protocol tips
Add calcein AM: 2 μM; EthD-1: 4 μM for 40 minutes at room temperature

Publication protocol

rTDSCs were seeded in 6-well plate (4×103 cells/well) and osteogenic-cultured in the designed oxygen tension conditions. On days 1, 4 and 7, cell viability of rTDSCs was analyzed with a LIVE/DEAD Viability/Cytotoxicity Assay Kit (Invitrogen) according to the instructions. Briefly, after washing with PBS for 2-3 times, rTDSCs in hypoxic and nomorxic cultures were incubated with fluorescent working solution (calcein AM: 2 μM; EthD-1: 4 μM) for 40 minutes at room temperature. Then, the live or dead rTDSCs in each group were viewed under a fluorescence microscopy (Olympus IX71). Quantification of cell viability was performed using the Image-Pro Plus software (Version 5.1, Media Cybernetics, Inc.)



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