Live/Dead Cell Double Staining Kit

Protocol tips

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	Add 100 µl of assay solution to cells and incubate the mixture at 37 ºC for 15 min.

Protocol tips
Add 100 µl of assay solution to cells and incubate the mixture at 37 ºC for 15 min.

Publication protocol

Cultured MCF-7 cells derived from human
metastatic breast carcinoma were grown in
DMEM medium, supplemented with 10 % FCS
(Sigma-Aldrich) in a 75 cm tissue culture flask (Corning) and maintained in a humidified incubator at 5 % CO2 and 37°C. Tumor cells were counted manually on hemacytometers. Cell viability was measured using a dye exclusion method (Invitrogen Corp.) where for each experiment; the cell viability
showed ≥ 90 % healthy cells after detachment
from culture flask and washing steps. The tumor
cells were stained with The “Live/Dead Cell Double Staining Kit” (Sigma-Aldrich), which is utilized for simultaneous fluorescence staining of viable and dead cells. This kit contains calceinAM and propidium iodide (PI) solutions, which stain viable and dead cells, respectively. The stained cells were serially diluted in phosphate buffered saline (PBS) to the desired
concentration for device testing. For recovery testing, the cells were diluted to between 10 and 10^3 cells/ml to minimize dilution and counting errors the cell counts were repeated 10 times. Blood samples were drawn from the free-cancer healthy donors into EDTA tubes and processed within 24 hours.

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