CD133 expression at the luminal surface of some epithelial tumour glands with shedding into the lumina within tumour spheroids. (A) CD133 (yellow) and EBP50 (magenta) were visualized within CT320X6 (upper panel) and HT29 (lower panel) spheroids by immunofluorescence staining. Spheroids were counterstained with TOPRO-3 (cyan). CD133 was found into the lumina and colocalized with EBP50 at the luminal surface of some acini. Acquisitions were taken at 70 μm depth (scale bar = 50 μm). (B) Dissociated cells from CT320X6 (upper histogram) and HT29 (lower histogram) spheroids were similarly labelled against CD133 and analysed by flow cytometry.