Objective Subcutaneous transplantation Lewis lung carcinoma model is commonly used in experimental studies. Researchers often choose different transplantation sites to create the models while little attention was paid on the effect of different inoculation sites on the formation of transplanted tumors. The aim of this study was to compare the effect of tumor cell inoculation at different sites on tumor formation in mice. Methods Lewis lung adenocarcinoma (ll2-luc-m38) cells stably expressing luciferase protein were subcutaneously injected into C57 BL/6 mice at the right armpit, right groin, or footpad, respectively. An IVIS spectrum in vivo imaging system was used to observe the tumor and metastasis formation. The survival time and mortality were recorded. H-E stained pathology was performed to examine the histological changes of the lung tissues and tumor metastesis. Results The tumor formation time was earlier in the armpit and groin groups, both with a tumor formation rate of 100%, while the tumors occurred later, with a tumor formation rate of 33% in the footpad group. The pulmonary metastasis rate was 70% in the groin group, 50% in the ampit group, and 0% in the footpad group, at the 21st day after inoculation. The footpad group had a high mortality. The tumors in the groin group and armpit group can be surgically resected, with a postoperative survival rate of 100%. Conclusions In this mouse model of subcutaneously transplanted Lewis adenocarcinoma, the groin and ampit groups have advantages such as a high tumor formation rate, good tolerance of tumor resection, low surgical mortality rate, easy to monitor, simple operation and high reproducibility. The axillary group has an even higher metastasis rate.