Figure 3.

SW620 metastatic cells exhibit an increased capacity for re-grow after 5FU treatment under independent and anchorage-dependent conditions compared to SW480 and RKO cell lines derived from primary tumors. (A) Experimental scheme to evaluate the ability of CRC spheroid cells to re-grow after 5FU treatment in anchorage-independent and-dependent conditions compared to cells from spheroids treated with vehicle (V) according to methodology. Spheroids treated were dissociated and seeded to 5000 cells/mL in low-attachment and Tissue culture-treated microplates, respectively. (B) Left panel: Photographs of cell cultures in low-attachment microplates on the 7th, 14th, and 21st day of incubation after spheroids dissociation and cell re-seeding (scale bar: 500 μm). Right panel: Cell viability of collected cells in these cultures was determined using AquaZombie dye on the 21st day of incubation. Fluorescence was measured by flow cytometry. (C) Colonies formed in high adherence surfaces (tissue culture-treated microplates) were stained with crystal violet on the 7th, 14th, and 21st days of growth after cell re-seeding. The colony formation efficiency (% CFE) post-5FU treatment was calculated on the 21st day. Images are representative of at least three independent experiments.