4.5. Subcutaneous Xenograft Models

Six nude mice were subcutaneously injected into the lateral backside with 1 × 10⁶ cells at their 180 passages. Cells were implanted into the right and left sides of each mouse. Calipers were used to measure the tumor volume, which was calculated using the formula Volume (π/6) × length × width². For Rapamycin experiment, when tumor volume reached 100 mm³, mice were intraperitoneally injected with Rapamycin (5 mg/kg) every other day with 5% Tween 80 plus 5% PEG400 as vehicle. Mice treated with Rapamycin did not show any sign of toxicity based on body weight, food/water intake, and activity. We measured the tumor volume twice per week for 3–4 weeks before we collected the tumors to measure their weight. When mice were euthanized, their tumors were removed and sectioned with hematoxylin and eosin (H&E) staining and immunostaining.