Axiovert 100 m inverted

Confocal fluorescence microscopy studies were performed with a Zeiss Axiovert 100-M inverted microscope equipped with a LSM 700 laser scanning unit and a 1.4 NA × 63 Plan-Apochromat oil-immersion objective. Cells were seeded in μ-Slide 8 well, ibiTreat, tissue culture treated, (Inter Instrument AS). To minimize photobleaching, laser power was typically 20% under maximum and the pinhole was set to 0.8–1.5. Multi-tracking was used for dual color imaging.

Endothelial cell spheroids were generated as previously described [55 (link)] and incubated with either IL-17A (supplied in platelet suspension buffer to correspond to the residual levels of the cytokine in the platelet supernatant following the stimulation of platelets; 5 ng/mL) or treated platelet supernatant as indicated. In vitro angiogenesis was quantified using an Axiocam MR digital camera with an Axiovert 100 M inverted microscope equipped with a Plan-NEOFLUAR objective (10×/0.30; Carl Zeiss, Jena, Germany). Quantification was performed by measuring the cumulative length of the sprouts that had grown out of each spheroid using a digital imaging software (Axioplan, Zeiss, Jena, Germany) and analysing 10 spheroids per group and experiment. The mean of the 10 spheroids per condition and per experiment was calculated.