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FIG. 1.
4D light-sheet fluorescence microscopy (LSFM) framework for zebrafish heart imaging and analysis. (a) Workflow of the zebrafish cardiac activity analysis, including major steps from zebrafish preparation in standard E3 medium with phenylthiourea (PTU) and tricaine, to 4D image acquisition, and to reconstruction and single cell tracking. (b) Simplified schematic illustration of the customized LSFM system construction. CL: cylindrical lens. EO: excitation objective. DO: detection objective. TL: tube lens. FL: filter. CAM: sCMOS camera. (c) Illustration of retrospective synchronization for 4D zebrafish image registration. Z-movie indicates a continuous image sequence at a certain depth along the z-axis. Each frame in the image sequence is represented by a red dot, and the starting and ending phases from end-diastole to end-systole are highlighted in the yellow box. (d) Procedures of cell segmentation and tracking. Raw, segmented, and successively registered images are presented from left to right. Individual cells are coded using pseudo-colors. (e) User-directed interaction, including cell selection and quantitative analysis, has been achieved in the virtual environment.