The multiscale real-time microscope integrated several features that facilitated rapid histological assessment of unsectioned tissue. First, the maximum field of view was adjustable. Second, the system incorporated a widefield white-light view that allowed rapid identification of inked margins and grossly apparent features, enabling the operator to rapidly locate margins on large specimens while avoiding areas that are not relevant to assessment. Finally, the authors implemented opportunistic averaging, a novel feature that enabled averaging without motion blur or other artifacts when translating the stage. All this allowed generation of mosaic images from data acquired by real-time user panning, enabling efficient review of procedures in a fraction of the time required to replay the data at real-time speed. Moreover, the framework presented here is consistent with pathology assessment performed using frozen section analysis, still a gold standard in histology. However, here it is done at much faster speeds and without freezing and cryo-sectioning, thus enabling real-time assessment of surgical specimens during surgery, which promises great reduction in the number of repeat surgeries, as is currently often needed due to postoperative histological findings.
Finally, the multiscale imaging approach demonstrated by the authors may be applicable to other less costly imaging modalities such as CFM, SIM, MUSE, or OCT that can perform high-speed imaging using multiple objectives or variable magnification with a single objective.
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