Figure 1

Establishment of hybrid spheroid formation conditions and histological examination of hybrid spheroid. (A) Method of hybrid spheroid formation. Canine MGT cells and DH82 were seeded in an ultra-low adhesion plate at a set ratio. Mixed cells were cultured to form an appropriate spherical shape and then used in experiments. The top panels (B–D) are CIPp and the bottom panels (E–G) are CIPm. (B,E) Hybrid spheroids shape according to culture time and ratio of canine MGT cells and macrophages. (C,F) Hybrid spheroids size according to culture time and ratio of canine MGT cells and macrophages. (D,G) H&E staining images of hybrid spheroid (canine MGT cell:DH82 = 3:1) sections (grey bar = 200 μm) and Confocal images of hybrid spheroids (red bar = 100 μm). Anti-CD206 (green) and anti-CD80 (red) were used to confirm polarization of DH82 (blue = nuclei DAPI staining). The results are presented as the mean ± SD of triplicate samples and are representative of 3 independent experiments. MGT, mammary gland tumor; H&E, hematoxylin, and eosin; DAPI, 4,6-diamidino-2-phenylindole.