Figure 5: Organization of cells within CT26 spheroids.
(a) Confocal image of Phalloidin staining on CT26 spheroids at the equatorial plane. Yellow dotted circle divides a spheroid into a high anisotropy (major axis is perpendicular to radius; aspect ratio of 2–2.5) and low anisotropy zone (aspect ratio ∼1.5). Scale bar, 50 μm. (b) Orientation of the major axis of cells at the equatorial plane. (c) Cell anisotropy has been estimated with the ratio of the major to the minor axis of the cell, as illustrated on the insert. Analysis of two axes has been performed using ImageJ and the fit ellipse function. Only cells with clearly marked cortical actin were used for the analysis. Data points (N=177) were grouped together in bins, with the error bar being the s.e.m. and the position being the average R/R0 position within the bin. The vertical line divides two zones: high anisotropy and low anisotropy zone. (d) Zoom-in on actin staining by the border of spheroids shows that cells at the surface are round and loosely bound, whereas after pressure is applied, the surface is well defined, and cells become more rectangular. Scale bar, 20 μm. (e) Bright-field/FITC image represents a spheroid with incorporated microbeads that are marked in green (FITC). Microbeads positioned at the surface of spheroids are highly deformed and represent a pear-like shape. Scale bars, 50 μm (bright field/FITC), and 20 μm (FITC).
