AIM: To observe the effects on human keratocytes by cationic liposome Lipofectamine^TM2000 (LF2000), to investigate the efficiency and safe range applied in human keratocytes, and establish basis for gene therapy of human keratocytes. METHODS: Human keratocytes cultured in vivo within 3 to 5 passages were used in experiment after being identified. The effects on proliferation of cultured human keratocytes by LF2000 with different concentrations and time were evaluated By MTT; the effects of LF2000 on the survival rate and its relation with 5,10,20,40,80mg/L concentration and time were detected by trypan blue staining. RESULTS: LF2000's effects on human keratocytes were related with concentration and time. The cellular proliferation and survival rate declined when concentration of LF2000 was above certain level, and this effect increased as time became longer. LF2000 had no effect with concentration under 40mg/L for 24 hours. CONCLUSION：LF2000 did not cause cytotoxicity during a concentration range“tested”, and it is hoped to play an important role in gene therapy of human keratocytes.