Chemotherapy-induced enrichment of DiI-retaining cells in vivo. (A) DiI-retaining tumor cells were enriched in vivo after treatment with 5-fluorouracil (FU). Negative: flow cytometry of CT-26 cells that were not stained with DiI: PBS-treated: flow cytometry of cells from tumor treated with phosphate-buffered saline (PBS); 5-FU-treated: flow cytometry of cells from tumor treated with 5-FU. (B) Tumor volume in mice after chemotherapy compared with the PBS-treated group (*P < 0.05, t-test). (C) Percentage of DiI-positive cells in tumors with or without chemotherapy (**P < 0.01, t-test). Error bars represent the standard deviation. DiI-labeled CT-26 cells were injected subcutaneously into Balb/C mice on day 1. When tumors grew to 10 × 10 mm, 5-FU 40 mg/kg was injected intraperitoneally every 3 days for a total of four injections. Vehicle-treated control mice were exposed to the same regimen, but injected only with PBS. Tumor volume was measured on days 7, 14, and 21. After the final treatment, tumors were digested and analyzed by flow cytometry on the next day. Experiments were repeated twice with similar results.