Ti e eclipse

Cell imaging was performed on a Zeiss Axiovert microscope equipped with a Cool Snap ES CCD camera (Ropper Scientific). Images were captured using 2×2 binning and 12 sequential z-planes were collected at 0.3-µm step intervals with an exposure time of 200 ms except for time-lapse video microscopy movies of Abp1–RFP and Sla1–RFP that were collected every second with five sequential z-planes (0.5 µm steps) and an exposure time of 100 ms.
For analysis of microtubules and vesicle motion, cell imaging was performed on a confocal spinning disk inverted microscope (Nikon TI-E Eclipse) equipped with a Yokogawa motorized confocal head CSUX1-A1 and an Evolve EMCCD camera. A dual color acquisition of six sequential z-planes (0.3-µm steps) was performed every second with an exposure time of 50 ms and 100 ms for GFP–Snc1 and Bik1–RFP, respectively. All image manipulations, montages, and fluorescence-intensity measurements were performed using ImageJ (Schneider et al., 2012 (link)). Tracking analysis and dot number quantifications were performed using Icy (de Chaumont et al., 2012 (link)).