To understand the antiproliferative efficacy of the combination treatments,
cells were analyzed for cell-cycle analysis at 1, 3, 5, or 8 days post treatment, as
described previously [10 (link)]. Briefly, 1 ×
106 cells were seeded in a 100-mm-diameter cell-culture dish (Becton
Dickinson, San Jose, CA). For 8-day treatment, the initial cell seeding was 0.5 ×
106 cells. Cells were allowed to attach for 24 h prior to treatment. At
different time points post treatment, cells were harvested by trypsinization and
centrifuged at 1300 rpm for 3 min at 4 °C (Sorvall Legend RT centrifuge, Thermo
Electron Corp., Waltham, MA). The supernatant was decanted, the cell pellet was washed
twice with ice-cold 1×DPBS (pH 7.4), and then the pellet was resuspended in a
solution (12.5 mg propidium iodide [Promega, Madison, WI], 250 mg sodium-citrate, and 250
μL Triton™ X-100 in 250 mL of water). The cells in the propidium iodide
solution were then incubated for 2 h in the dark in a cold room and then analyzed by flow
cytometry (FACScan flow cytometer, BD Biosciences, San Jose, CA). ModFit LT software
(Verity Software House, Inc., Topsham, ME) was used for data analysis.
cells were analyzed for cell-cycle analysis at 1, 3, 5, or 8 days post treatment, as
described previously [10 (link)]. Briefly, 1 ×
106 cells were seeded in a 100-mm-diameter cell-culture dish (Becton
Dickinson, San Jose, CA). For 8-day treatment, the initial cell seeding was 0.5 ×
106 cells. Cells were allowed to attach for 24 h prior to treatment. At
different time points post treatment, cells were harvested by trypsinization and
centrifuged at 1300 rpm for 3 min at 4 °C (Sorvall Legend RT centrifuge, Thermo
Electron Corp., Waltham, MA). The supernatant was decanted, the cell pellet was washed
twice with ice-cold 1×DPBS (pH 7.4), and then the pellet was resuspended in a
solution (12.5 mg propidium iodide [Promega, Madison, WI], 250 mg sodium-citrate, and 250
μL Triton™ X-100 in 250 mL of water). The cells in the propidium iodide
solution were then incubated for 2 h in the dark in a cold room and then analyzed by flow
cytometry (FACScan flow cytometer, BD Biosciences, San Jose, CA). ModFit LT software
(Verity Software House, Inc., Topsham, ME) was used for data analysis.
