I have no idea why my orthotopic mouse models have failed.

I have used gastric cancer cell lines, and I injected the cells into a mouse stomach.

Here is my procedure and a result photo as a failed example.

1. Detach gastric cancer cells with 0.25% trypsin, then wash the cells with PBS

2. Centrifuge for 3 min, 1200 rpm at room temperature

3. Suction the trypsin and PBS, then put 1 mL of PBS

4. Get 1 x 10^8 cells in 1 mL of PBS, which is an example, counted by a cell counter

5. Transfer 100 uL of cell suspension to get 5 x 10^6 cells for 2 mice

4. Centrifuge for 0.5 min

5. Remove the PBS and put 15 uL of Matrigel and 15 uL of PBS

6. Suspend the 30 uL of gastric cancer cells and put it into a 0.5 mL insulin syringe

7. Keep the cells in the ice

8. Anesthetize a syngeneic mouse (4-6 weeks old) with 2% Isoflurane

9. Open the abdominal cavity

10. Take out the stomach then inject the cells, and try to inject them into the serosa; however, I sometimes inject the cells into the muscle layer.

11. If bleeding occurs after cell injection, the injection has failed

12. Suture the peritoneal cavity

13. Remove the stomach after a month, and dissect the stomach

Mice have never died, and they do not have any cancer organs even if I injected gastric cancer cell lines in their stomach.

Metastasis has also never been discovered.

This experiment has been conducted with the naked eye without a microscope.

I have been frustrated. What is my problem? Would you do me a favor, please?