The aim of this work is to optimize conditions of malignant cells cultivation in soft agar for subsequent immunohistochemical analysis of formed three-dimensional colonies. Methods. Cultivation of breast carcinoma cell line MCF-7 in soft agar, immunohistochemical and immunofluorescence detection of epithelial antigen and mTOR kinase in cultured cells. Results. We describe a methodical approach to the cultivation of cells in soft agar, which allows to carry out morphological, morphometric and immunochemical analysis of the studied cells. Conclusions. The proposed method provides an additional characterization of cells growing in soft agar, which will be useful in basic research and in evaluation of the effectiveness of anticancer drugs

Keywords: soft agar, evaluation of colony formation, three-dimensional culture of malignant cells