ZEISS Airyscan: Optimizing Usage for Fast, Gentle, Super-Resolution Imaging.

The Zeiss Airyscan microscope transforms a diffraction-limited, point-scanning confocal microscope into a super-resolution microscope using a specialized 32-channel Airyscan detector. By improving resolution twofold and signal-to-noise ratio eightfold relative to conventional confocal microscopes while retaining confocal functionality, the Airyscan microscope has become a very popular super-resolution imaging tool for cell biologists. In this chapter, we describe the fundamentals of Airyscan imaging, with the aim of helping the reader determine the proper acquisition settings for different types of experiments, optimize imaging conditions, and process the raw Airyscan images to obtain final images with the best quality. We also provide some tips, tricks, and best practices for Airyscan imaging. Of note, while our focus is on the Airyscan function of this microscope rather than its conventional confocal function, the Airyscan unit comes as an add-on to the conventional Zeiss laser scanning confocal microscope. This protocol is for the first generation Airyscan Zeiss 800 series microscope.