Eclipse t100 inverted microscope

To allow spheroid formation, 3.0 × 10⁵ colonic MSCs or colon cancer MSCs were transferred in 0,5 ml serum-free medium [55 (link)] in polypropylene 15 ml conical tubes (Falcon, BD) and incubated with or without CRC exosomes in 1:1000 (exosome mg/cell number) ratio until 72 hours at 37°C and 5% CO₂ in continuous rotation. Then cell spheroids were transferred in chamber slides (Falcon, BD), fixed in 3% paraformaldehyde, washed and analysed by a Nikon Eclipse T100 inverted microscope (Nikon Instruments Inc.) or by CLSM. The volume of the cMSCs spheroid structures were calculated using the following formula: (4π/3)xa²c, where a is the equatorial radius of the spheroid, c is the distance from center to pole along the symmetry axis. The pH of cMSCs spheroids supernatant was estimated by the use of a pH 123 Microprocessor pH Meter (Hanna Instruments, Italy).

Images were acquired with a Nikon Eclipse T100 inverted microscope (Nikon Instruments Inc., Melville, NY, United States) equipped with an LWD 10X 0.40 N.A. phase contrast objective, a Nikon DS-Fi1 color camera, and the NIS-Elements F v3.0 software (Nikon Instruments Inc.).