Fig. 1

Characterization of 3D HNSCC spheroids. a Schematic representation of cell seeding used for the formation of F5, F5M2 and F5M5 spheroids. b, c Typical microscopy images of cryosections of homo- (F5) and heterospheroids (F5M2 and F5M5) at day 5 post-seeding, stained with (b) DAPI nuclear dye, PKH67 and E-Cadherin markers and (c) vimentin and collagen markers. MeWo cells were pre-stained with PKH67 membrane dye. d Colocalization of vimentin and MeWo cells, stained with PKH67, within co-culture spheroids. Scale bar – 100 µm. e The percentage of MeWo cells in F5M2 ( ) and F5M5 ( ) co-culture spheroids 5 days post-seeding. The graph represents the mean ± SD (box) together with minimum and maximum limits (whiskers) [n = 87; ***p < 0.001, using two-sample t-test]. f Semi-quantitative image analysis of collagen expression in F5 ( ), F5M2 ( ), and F5M5 ( ) spheroids. Data presented as mean ± SD [n = 3–5; *p < 0.05; **p < 0.01, using ANOVA]