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Deconvolution-free Subcellular Imaging with Axially Swept Light Sheet Microscopy

2015; Elsevier BV; Volume: 108; Issue: 12 Linguagem: Inglês

10.1016/j.bpj.2015.05.013

1542-0086

Kevin M. Dean, Philippe Roudot, Erik S. Welf, Gaudenz Danuser, Reto Fiolka,

Cell Image Analysis Techniques

The use of propagation invariant Bessel beams has enabled high-resolution subcellular light sheet fluorescence microscopy. However, the energy within the concentric side lobe structure of Bessel beams increases significantly with propagation length, generating unwanted out-of-focus fluorescence that enforces practical limits on the imaging field of view size. Here, we present a light sheet fluorescence microscope that achieves 390 nm isotropic resolution and high optical sectioning strength (i.e., out-of-focus blur is strongly suppressed) over large field of views, without the need for structured illumination or deconvolution-based postprocessing. We demonstrate simultaneous dual-color, high-contrast, and high-dynamic-range time-lapse imaging of migrating cells in complex three-dimensional microenvironments, three-dimensional tracking of clathrin-coated pits, and long-term imaging spanning >10 h and encompassing >2600 time points.