Axio observer z 1 sd inverted microscope

Live-cell imaging experiments were performed essentially as previously described [34 (link)]. Briefly, MCF-7 or NCI-H460 cells were seeded onto LabTek II chambered cover glass (Nunc, Penfield, NY, USA) containing 1 mL of RPMI supplemented with 5% FBS, and incubated overnight at 37 °C under 5% CO₂. Then, the medium was replaced with 1 mL of RPMI without phenol red supplemented with 5% FBS, in the presence of 12 μM of the compound. Images were captured at 10 min intervals for 48 h under differential interference contrast (DIC) optics, with a 63x objective on an Axio Observer Z.1 SD inverted microscope (Carl Zeiss), equipped with an incubation chamber with the temperature set to 37 °C and an atmosphere of 5% CO₂. Movies were generated from the time-lapse images using ImageJ software (version 1.44, Rasband, W.S., ImageJ, U. S. National Institutes of Health, Bethesda, MD, USA). The number of cells that were arrested at mitosis, apoptotic, or bypassed cytokinesis was scored.

For live-cell imaging, a total of 0.09 × 10⁶ A549 cells were seeded into a LabTek II chambered cover glass (Nunc, Penfield, NY, USA) in complete RPMI culture medium. Sterile water was added to the remaining wells to guarantee a humidified atmosphere. The cells were incubated overnight at 37 °C under 5% CO₂. Then, the medium was replaced by fresh medium in the presence of MPS-1/CENPE inhibitors alone or in combination with navitoclax at the concentration of the respective synergistic points. Time-lapse images were taken every 5 min over a 48 h period using differential interference contrast (DIC) optics, and a 63× objective on an Axio Observer Z.1 SD inverted microscope (Carl Zeiss, Oberkochen, Germany). The microscope is equipped with an incubation chamber set to 37 °C and 5% CO₂. ImageJ software (version 1.47, Rasband, W.S., ImageJ, U. S. National Institutes of Health, Bethesda, MD, USA) was used to create movies from the time-lapse images.