The growth of malignant tumors occurs in 3D environments and depends on the presence of a stromal component, which performs critical functions of tumor-cell protection and growth support. Therefore, the development and analysis of tumor models in 3D cell cultures in vitro, including coculture systems, hold significant interest. In this study, the results of 3D culturing of two human melanoma cell lines using the hanging drop method with and without human mesenchymal stem cells (MSCs) have been presented. The melanoma lines have been shown to behave differently in 3D cultures. In particular, the Mel Cher melanoma cells are capable of the formation of uniform spheroids within 24 h, whereas the MeWo cells failed to form spheroids even after 2 days of culture under similar conditions. However, in both cases, the coculturing of the melanoma cells with MSCs resulted in the formation of compact 3D cell spheroids. The visualization of MeWo cells and MSCs in mixed spheroids using fluorescent dyes revealed a certain clustering of the melanoma cells. The observed properties of the melanoma cells in homogeneous and heterogeneous spheroids may be used in the complex analysis of antimelanoma chemotherapy drugs and the estimation of their therapeutic properties.