Abstract: This article describes the preparation of the N-tert-butyl-α-phenylnitrone (PBN) liposomes and their related characteristics. The PBN liposomes were prepared by film dispersion-supersonic method and the formula of liposomes was optimized by orthogonal uniform design. RP-HPLC was used to qualify the amount of PBN that entered into the hepatoma cells. Necrosis rate was also investigated by fluorescence activated cell sorter (FACS) after PBN liposomes transfection. Result showed that the mean particle size, entrapment efficiency, and polydispersity of the resulting PBN-liposome were 137.5 nm, 71.52% and 0.286, respectively. PBN liposomes can enter into the tumor cell stably and they have higher affinity to hepatoma cell compared with free PBN resulting in a higher necrosis rate after transfection. These results provide a potential method for early diagnosis and treatment of cancer using specific spin trapping probe targeting tumor cells.