Combined Multi-Plane Tomographic Phase Retrieval and Stochastic Optical Fluctuation Imaging for 4D Cell Microscopy

摘要

Super-resolution fluorescence microscopy provides unprecedented insight intocellular and subcellular structures. However, going beyond the diffractionbarrier comes at a price since most far-field super-resolution imagingtechniques trade temporal for spatial super-resolution. We propose thecombination of a novel label-free white light quantitative phase tomographywith fluorescence imaging to provide high-speed imaging and spatialsuper-resolution. The non-iterative phase reconstruction relies on theacquisition of a single image at each z-location and thus enablesstraightforward 3D phase imaging using a classical microscope. We realizedmulti-plane imaging using a customized prism for a simultaneous acquisition of8 planes. This allowed us to not only image live cells in 3D at up to 200 Hz,but also to integrate fluorescence super-resolution optical fluctuation imagingwithin the same optical instrument. This 4D microscope platform unifies thesensitivity and high temporal resolution of phase tomography with thespecificity and high spatial resolution of fluorescence imaging.