Figure 1.

Sphere formation abilities of Gprc5a^−/− Kras-mutant LUAD cells. Single cell suspensions of Gprc5a^−/− MDA-F471 cells (see Materials and Methods) were embedded in Matrigel™ and plated at the rim of each well. Media containing 5% FBS was added in the middle of each well. Cells were incubated for 6–7 days until they formed spheres. Sphere forming units were calculated as percentages of the number of formed spheres relative to the number of seeded cells. To assess the self-renewal ability of sphere forming cells, spheres were propagated for up to five generations. (A) Sphere forming units (SFUs) of MDA-F471 cells across five generations. Error bars represent standard deviations between technical triplicates of each of three independent experiments. (B) Representative bright-field images of MDA-F471 G1-G5 spheres visualized by Axiovert inverted microscope at 10X magnification and analyzed by Carl Zeiss Zen 2 image software. Scale bar = 100 μm. (C) Scatter plots of the diameters (μm) of 30 spheres from G1 up to G5 for MDA-F471. Data were reported as mean ± SD (P < 0.0001 by one-way ANOVA).