Three-Color Simultaneous Live Imaging of Autophagy-Related Structures
Simultaneous live cell imaging of multiple proteins helps to analyze mobility and interactions among proteins over time. Since autophagosomes depend on other organelles for their formation, it is necessary to observe this process with multiple fluorsphores to mark multiple organelles and the autophagosomes. To do so, we set up three cameras on one microscope to be able to acquire three colors at the same time. Here we describe the setup using a Yokogawa spinning disk confocal microscope (CSU-W1) with Andor TuCam system attaching 3 × Zyla 4.2 CMOS cameras (Andor) and detail the method for acquiring live images.