Purpose To compare and investigate the oncogenic and metastatic phenotypes in nude mice by injection of rat hepatocellu-lar carcinoma ( HCC) cells L2 via two different routes. Methods Twenty of 7-week-old female BALB/cA mice were randomly divided into 2 groups. After the injections of L2 from liver or spleen lobe, the survival rate, tumor formation rate, carcinogenic features, and metastasis were comparatively studied. Results All of the liver orthotopic nude mice developed liver cancer (100%) with 60% lung metastasis rate, and exhibited an expansive growing pattern with surrounding invasion. In the spleen orthotopic model, 78% mice de-veloped HCC in spleen, with 67% liver metastatic rate and 11% lung metastatic rate, lower than the liver orthotopic model ( P <0. 05). But the microscopically hilar lymph node metastasis rate was 33%. Conclusion The direct liver injection of L2 in female nude mice is a better modeling method for studying the mechanism of both carcinogenesis and metastasis, as well as the evaluation of therapeutic effect of liver cancer.