AbstractStimulated Emission Depletion (STED) nanoscopy has become one of the most used nanoscopy techniques over the last decade. However, most recordings are done in specimen regions no larger than 10–30 × 10–30 µm2 due to aberrations, instability and manual mechanical stages. Here, we demonstrate automated STED nanoscopy of extended sample regions up to 0.5 × 0.5 mm2 by using a back-aperture-stationary beam scanning system. The setup allows up to 80–100 x 80–100 µm2 field of view (FOV) with uniform spatial resolution, a mechanical stage allowing sequential tiling to record larger sample areas, and a feedback system keeping the sample in focus at all times. Taken together, this allows automated recording of theoretically unlimited-sized sample areas and volumes, without compromising the achievable spatial resolution and image quality.
Tuning donut profile for spatial resolution in stimulated emission depletion microscopy
