LIVE/DEAD® Cell Imaging Kit (Life technologies) was used to quantify cell viability. The ‘Live’ and ‘Dead’ parts of the stain were mixed following the manufacturer protocol. The mixed solution was diluted 3 times with cell media. This 3x-diluted solution was mixed with cell samples at 50/50 volume ratio to stain the cells. NucBlue® Live ReadyProbes® Reagent (Life technologies) were used to identify cell nuclei to accurately quantify cell counts.
Apoptosis was detected using Alexa Fuor® 488 Annexin V (Thermo Fisher Scientific). The dilution of cell samples and Dox were done with 1X Annexin- binding buffer (Thermo Fisher Scientific). Dox was mixed with the cell samples as described for other drug assays at a concentration of 10 μM. 25 μl of Alexa Fuor® 488 Annexin V (Thermo Fisher Scientific) was added to 100 μl cell-drug samples as an early-stage apoptosis marker. The cell sample with the Alexa Fuor® 488 Annexin V was incubated at room temperature for 15 min, loaded to the microfluidic device and then incubated at 37 °C in 5% CO2 for 45 mins. Fluorescence images (FITC and TRITC) were taken at time points of 1, 3, 6 and 9 hrs. In between imaging, the devices were incubated at 37 °C in 5% CO2.
Apoptosis was detected using Alexa Fuor® 488 Annexin V (Thermo Fisher Scientific). The dilution of cell samples and Dox were done with 1X Annexin- binding buffer (Thermo Fisher Scientific). Dox was mixed with the cell samples as described for other drug assays at a concentration of 10 μM. 25 μl of Alexa Fuor® 488 Annexin V (Thermo Fisher Scientific) was added to 100 μl cell-drug samples as an early-stage apoptosis marker. The cell sample with the Alexa Fuor® 488 Annexin V was incubated at room temperature for 15 min, loaded to the microfluidic device and then incubated at 37 °C in 5% CO2 for 45 mins. Fluorescence images (FITC and TRITC) were taken at time points of 1, 3, 6 and 9 hrs. In between imaging, the devices were incubated at 37 °C in 5% CO2.