Focus image scanning microscopy for sharp and gentle super-resolved microscopy
To date, the feasibility of super-resolution microscopy for imaging live and thick samples is still limited. Stimulated emission depletion (STED) microscopy requires high-intensity illumination to achieve sub-diffraction resolution, potentially introducing photodamage to live specimens. Moreover, the out-of-focus background may degrade the signal stemming from the focal plane. Here, we propose a new method to mitigate these limitations without drawbacks. First, we enhance a STED microscope with a detector array, enabling image scanning microscopy (ISM). Therefore, we implement STED-ISM, a method that exploits the working principle of ISM to reduce the depletion intensity and achieve a target resolution. Later, we develop Focus-ISM, a strategy to improve the optical sectioning and remove the background of any ISM-based imaging technique, with or without a STED beam. The proposed approach requires minimal architectural changes to a conventional microscope but provides substantial advantages for live and thick sample imaging.
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Tortarolo, G., Zunino, A., Fersini, F., Castello, M., Piazza, S., Sheppard, C. J. R., Bianchini, P., Diaspro, A., Koho, S., & Vicidomini, G. (2022). Focus image scanning microscopy for sharp and gentle super-resolved microscopy. Nature Communications, 13(1), Article 7723. https://doi.org/10.1038/s41467-022-35333-y
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