CASP3 and cancer: To better understand presence of apoptosis and hypoxia, we stained the apoptosis-induced cleaved Caspase-3 (active Caspase-3) and the hypoxia-marker CA9 together with cytokeratin and the fibroblast marker MRC2 in 12-day-old heterospheroids of HT-29 and SW480 cancer cells using 4-plex immunofluorescence (Figure 5B).