Objective To establish and characterize patient ⁃derived breast cancer organoids ，and to explore their potential role as promising preclinical models for individualized chemotherapeutic drug sensitivity assays. Methods Breast cancer tissues（different molecular subtypes）of 4 female patients who underwent radical mastectomy were obtained. Freshly resected tumors were collected and cells were isolated. Isolated cells were plated in Matrigel drops and overlaid with organoid culture medium for three ⁃dimensional culture. Tissues and organoids were fixed，followed by dehydration，paraffin embedding，and serial sectioning. Tissues and organoids sections were subjected to H&E and immunohis⁃tochemical staining（ER，PR，HER2，and Ki⁃67）in order to investigate whether the histological characteristics of the parental tumors were maintained in the breast cancer organoids. After treatment of breast cancer organoids with different concentrations of chemotherapeutic drugs for 5 days，cell viability was determined using the CellTiter⁃Glo® 3D reagent and drug sensitivities were analyzed. Results Breast cancer organoids recapitulated the histological features and marker expressions of their parental tumors. And the sensitivity of organoid chemotherapeutic drugs of different molecular subtypes in breast cancer was different. Conclusion The in vitro culture system of breast cancer organoids has been successfully established in this study，which can reflect the distinct molecular subtypes and histological features of the parent tumor，and can be used for in vitro chemotherapy drug test. These data suggest that breast cancer organoids may be an excellent preclinical model for studying clinical response to chemotherapeutic drugs in a personalized way，as well as for providing reference for clinic drug use.