Figure 4.

Increased tumor growth of stem‐like ECE1c^K6R cells in vivo. (A) DLD‐1 clone cells expressing either Flag‐tagged ECE1c^WT or ECE1c^K6R proteins (or Mock cells) were grown for 21 days in soft agar, and anchorage‐independent growth was determined. Representative images of crystal violet‐stained colonies obtained at a magnification of 40×. (B) Colonies formed from cells in A were counted and plotted as total colonies/well. (C) Sizes of colonies (μm in diameter) formed in A were measured using micrometrics se premium 4 software. (D) Cells (2 × 10⁶) were subcutaneously injected into NOD/SCID mice, and tumor volume (mm³) was monitored. After the mice were euthanized on day 22, tumors were photographed, measured, and fixed in paraffin. (E) Paraffin slices of tumors from the experiment in D were analyzed by IHC with anti‐FLAG (left) and anti‐Ki‐67 (right) antibodies (scale bars upper panels = 150 µm; lower panels = 50 µm). Data represent average ± SEM (n = 3). ANOVA and Tukey tests were used. **P ≤ 0.01, ***P ≤ 0.001.