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- W2513303908 abstract "Anisotropy imaging is a powerful tool to study the organization, composition and dynamics of molecules in biological systems. Changes in the organization and conformation of biomolecules within cells and tissues can be visualized at the single pixel level. Fluorescence anisotropy is typically probed with laser scanning and epifluorescence-based techniques. However, those techniques are limited in either axial resolution, image acquisition speed, or, by photobleaching. In the last decade, selective plane illumination microscopy (SPIM) has emerged as the best choice for three-dimensional time lapse imaging since it combines axial sectioning capability with fast, camera-based image acquisition, and minimal light exposure [1,2]. We demonstrate how SPIM can be used for three-dimensional fluorescence anisotropy imaging of live cells [3]. To verify the accuracy of our SPIM system, we first measured and compared the anisotropy of Rhodamine110 solutions with different water/glycerol ratios with results obtained using a spectrofluorimeter. Then, we visualized the aggregation states of different fusion proteins in live cells. Further, we were able to show by 3D time lapse anisotropy imaging that formation of membrane protrusions and paxillin aggregates precedes focal adhesion assembly.Work supported in part by NIH grants P50 GM076516 and P41 GM103540.[1] Huisken, J. et al., Optical Sectioning Deep Inside Live Embryos by Selective Plane Illumination Microscopy. Science 305, 1007-1009 (2004).[2] Hedde, P.N. et al., Rapid Measurement of Molecular Transport and Interaction inside Living Cells Using Single Plane Illumination. Sci. Rep. 4, 7048 (2014).[3] Hedde, P.N. et al., 3D fluorescence anisotropy imaging using selective plane illumination microscopy. Opt. Express 23, 22308-22317 (2015)." @default.
- W2513303908 created "2016-09-16" @default.
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- W2513303908 date "2016-02-01" @default.
- W2513303908 modified "2023-09-26" @default.
- W2513303908 title "Fluorescence Anisotropy Imaging in 3D with Single Plane Illumination Microscopy" @default.
- W2513303908 doi "https://doi.org/10.1016/j.bpj.2015.11.2578" @default.
- W2513303908 hasPublicationYear "2016" @default.
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