Figure 3
CSCs phenotypes in the canine PAC cell lines. (a, b) Sphere-formation abilities were assessed in low-attachment culture conditions. Cells were seeded at 2000 cells/well, and the number of spheres were counted visually under a light microscope at 40 × magnification after 10 days. Representative images of spheres in CPACd-AB4 and CPACd-CG11 are shown in (a). Scale bar: 500 µm. Data are represented as the mean ± SD. Results are representative of three independent experiments. (c) SP populations in each cell lines were assessed using Vybrant DyeCycle Violet (DCV). Cells (5 × 105) were labeled with 5 µM DCV alone or in combination with 50 µM verapamil. SP populations were detected using flow cytometer with violet filter (450/40 and 660/20). Results are represented as the mean ± SD of three independent experiments. *P < 0.05, **P < 0.01: The number of spheres and SP populations are significantly higher than that of the parent cell lines; ††P < 0.01: The number of spheres is significantly lower than that of the parent cell lines.
