Beyond the Diffraction Limit by LightSheet Microscopy
Bi-Chang Chen1*
1Research Center for Applied Sciences, Academia Sinica, Taipei, Taiwan
* Presenter:Bi-Chang Chen,
By combining the intrinsic optical sectioning with wide-field detection, lightsheet microscopy allows fast imaging speeds to record multi-megapixel imaging of the selected plane in a single exposure. In order to break diffraction limit, coupled with structural illumination, we are now able to achieve final lateral and axial resolutions of 130 nm and 200 nm, respectively. On the other hand, one could combine the advantage of localization microscopy and lightsheet microscopy to have super-resolved cellular or deep-tissue imaging in 3D across large field of view. With densely labeled spontaneous blinking fluorophore, these allow us to construct 3D super-resolution multi-cellular imaging at high speed (~minutes) by lightsheet localization microscopy. Moreover, expansion microscopy (ExM) was invented to detour the optical diffraction limit by physically expanding the samples to ~4 times larger than original with swellable polymer and lightsheet microscopy enables imaging such large specimens rapidly at high spatial and temporal resolution. We applied various lightsheet microscopies to image systems spanning five orders of magnitude in space and time, including the single molecule localization, mitochondria dynamics within division apparatus, nanoscopic Dopaminergic neurons distribution across an intact Drosophila brain and blood vessel distribution in a whole mouse brain.

Keywords: lightsheet microscopy , superresolution , fluorescence imaging , 3D imaging