Objective: To construct a UV-responsive human epidermal growth factor receptor 2 (HER-2)-targeting immunoliposome, and investigate the improvement of chemotherapy efficacy on breast cancer. Methods: We fabricated the trastuzumab Fab fragment-decorated immunoliposome by thin film-dispersion method, which formed intermolecular cross-linking through the diacetylenic group by ultra-violet (UV) irradiation. The liposomal morphology was observed with transmission electron microscope. The particle size distribution and serum stability were evaluated by dynamic light scattering. Flow cytometry was performed to compare the intracellular uptake of liposomal and free drugs. The in vitro and in vivo anti-tumor efficacy of liposomal drugs against MCF-7 cells were evaluated by CCK-8 and subcutaneous tumor inhibition analysis, respectively. Results： After UV irradiation, the immunoliposome exhibited a fine spherical structure, better serum stability and slower drug release with a decreased mean diameter of (88.1±13.2) nm. The cellular uptake of immunoliposome(PC-DOX-Fab) was significantly enhanced comparing with non-targeting liposomes(PC-DOX-BSA) and free drug(DOX)(P<0.05). Cytotoxicity assays against HER-2+ cancer cells showed that the tumor suppressing efficacy of immunoliposomal drugs was significantly improved comparing with PC-DOX-BSA and free DOX at the same condition(P<0.05). In vivo studies demonstrated that the treatment of PC-DOX-Fab significantly reduced the tumor growth compared with that of PC-DOX-BSA and free DOX (P<0.05). Conclusion: In this study, we identified a novel liposomal drug delivery system for improved chemotherapy against HER-2-positive breast cancer cells, which owned a fine spherical structure and excellent serum stability. The in vitro and in vivo experimental results clearly suggested that this UV-responsive immunoliposome exhibits potent tumor suppressing activities, which deserved further investigation for clinical application.