A microscope-based method for quantitative analysis of contact-dependent cell-to-cell molecular communication and transfer between cells, the method comprising the steps: (i) obtaining of first singularized cells containing or expressing one or more traceable substances to provide donor cells; (ii) obtaining of second singularized cells not containing or expressing said one or more traceable substances to provide acceptor cells; (iii) combining said donor and acceptor cells in a ratio so that a majority of donor cells are surrounded by acceptor cells when grown in co-culture on a substrate and that the donor cells are essentially more than 200 micrometers apart from each other to obtain neighborhood regions wherein a contact-dependent cell-to-cell substance transfer can take place between said one donor cell and its surrounding acceptor cells, (iv) culturing of said donor cells and acceptor cells in a culture medium in the presence of a substrate for a first period of time to allow donor and acceptor cells to attach to said substrate; (v) replacing said culture medium by a medium wherein the proliferation of cells is inhibited and, optionally, washing cells attached to said substrate; (vi) co-culturing of said donor and acceptor cells in said proliferation inhibiting medium for a second period of time to allow for contact-dependent cell-to-cell transfer between said donor and acceptor cells; (vii) imaging of donor and acceptor cells to identify single donor cells which are more than 200 micrometers apart from the next donor cell but in contact to surrounding acceptor cells and thereby defining by microscopy a region of interest containing in a space volume the identified single donor cell and a number of surrounding acceptor cells in contact relationship: (viii) determining the amounts of traceable substance contained within the volume picture elements (volumetric pixel or voxel) of said region of interest and defining the volume picture elements which are located within the cell boundaries of said donor and acceptor cells by cell segmentation; and (ix) calculating the amount of traceable substance present within the volume elements located within the acceptor cell(s) surrounding the single donor cell to determine the amount of contact-dependent cell-to-cell substance transfer between said donor cell(s) and surrounding acceptor cell(s). (i) obtaining of first singularized cells containing or expressing one or more traceable substances to provide donor cells; (ii) obtaining of second singularized cells not containing or expressing said one or more traceable substances to provide acceptor cells; (iii) combining said donor and acceptor cells in a ratio so that a majority of donor cells are surrounded by acceptor cells when grown in co-culture on a substrate and that the donor cells are essentially more than 200 micrometers apart from each other to obtain neighborhood regions wherein a contact-dependent cell-to-cell substance transfer can take place between said one donor cell and its surrounding acceptor cells, (iv) culturing of said donor cells and acceptor cells in a culture medium in the presence of a substrate for a first period of time to allow donor and acceptor cells to attach to said substrate; (v) replacing said culture medium by a medium wherein the proliferation of cells is inhibited and, optionally, washing cells attached to said substrate; (vi) co-culturing of said donor and acceptor cells in said proliferation inhibiting medium for a second period of time to allow for contact-dependent cell-to-cell transfer between said donor and acceptor cells; (vii) imaging of donor and acceptor cells to identify single donor cells which are more than 200 micrometers apart from the next donor cell but in contact to surrounding acceptor cells and thereby defining by microscopy a region of interest containing in a space volume the identified single donor cell and a number of surrounding acceptor cells in contact relationship: (viii) determining the amounts of traceable substance contained within the volume picture elements (volumetric pixel or voxel) of said region of interest and defining the volume picture elements which are located within the cell boundaries of said donor and acceptor cells by cell segmentation; and (ix) calculating the amount of traceable substance present within the volume elements located within the acceptor cell(s) surrounding the single donor cell to determine the amount of contact-dependent cell-to-cell substance transfer between said donor cell(s) and surrounding acceptor cell(s).