Fig. (4) A robust algorithm can capture both local (single plane) data and global (whole spheroid) phenomena. Using a specific field within a well of a 96-well plate an algorithm for analysis of CK5Pro-GFP expression in MCTS can be used to capture data in a single plane or globally by using maximal projections. Panels 1-4 (bottom right) show the maximal projection of 8 Z-planes. Panel 1 shows the stained nuclei (Hoechst, blue) and the CK5Pro-GFP expression (green). Panel 2 shows the regions of the image tiled with spots (arbitrary colors). Panel 3 shows spots selected by fluorescence intensity and size (selected spots in green, rejected spots in red). Panel 4 shows the entire spheroid region across which high-content parameters were summed.