Background. The development of breast cancer (BCa) is largely determined by the characteristics of the tumor microenvironment (ТМЕ), which undergoes significant changes during the progression of the disease. Mast cells (MCs) are among the least studied components of the TME. The aim of the work was to investigate the relationship between the density of infiltration and the functional activity of MCs with indicators of osteopontin (OP) expression in BCa tissue. Materials and Methods. The study was conducted on the postoperative material of 15 patients with fibroadenoma and 78 patients with stage I—II BCa. MCs in the tissue of benign and malignant breast tumors were detected by a histochemical method using toluidine blue. The functional activity of MCs was calculated by the degranulation index. The OP expression in tumor tissue was assessed by the immunohistochemical method. Results. The density of MCs infiltration and their functional activity are associated with such indicators of BCa malignancy as tumor size, lymph node involvement, tumor grade, molecular subtype, proliferative activity, and PR- and HER2/neu-expression status. A high expression of OP in the stromal component of BCa is associated with the growth of the tumoral MCs population, metastatic lesions in regional lymph nodes, and a low differentiation grade of the tumors. In addition, OP is involved in the regulation of MCs in the tissue of the luminal B and basal molecular BCa subtypes. The level of OP expression in the parenchymal component of BCa is associated with the number of infiltrated MCs in the presence of metastatic lesions of regional lymph nodes. Conclusions. The identified relationship of OP expression level with the topology and functional activity of MCs in BCa tissue, depending on the clinical status of patients, indicates the prospects for their use in predicting the aggressiveness of the tumor process.