Focus-ISM for Sharp and Gentle Super-Resolved Microscopy (Q6708399)

To date, the feasibility of super-resolution microscopy for imaging liveand thick samples is still limited. Stimulated emission depletion (STED)microscopy requires high-intensity illumination to achieve sub-diffractionresolution, potentially introducing photodamage to live specimens. Moreover, the out-of-focus background may degrade the signal stemming fromthe focal plane. Here, we propose a new method to mitigate these limitations without drawbacks. First, we enhanced a STED microscope witha detector array, enabling image scanning microscopy (ISM). Therefore,we implemented STED-ISM, a method that exploits the working principleof ISM to reduce the depletion intensity and achieve a target resolution.Later, we developed Focus-ISM, a strategy to improve the optical sectioning and remove the background of any ISM-based imaging technique, withor without a STED beam. The proposed approach requires minimal architectural changes to a conventional microscope but provides substantialadvantages for live and thick sample imaging.