Current paper presents the results of the chlorine e6 (Ce6) study on 2D and 3D models of FaDu cells culture. The 2D model or monolayer was used for investigation of Ce6 distribution within individual cells and their organelles. The 3D model or multicellular tumor spheroids were used for estimation of cells’ metabolic processes by the investigation of the Ce6 fluorescence distribution within spheroid's layers and Ce6 fluorescence lifetime. It was shown that 3D cell cultures and Сe6 allows estimating the cells’ metabolic processes better than in 2D monolayer cell cultures. Also, this model allows estimating the photodynamic effect depending on the proximity to the surface of different areas inside the heterogeneous 3D structure.