Combined Selective Plane Illumination Microscopy and FRAP Maps Intranuclear Diffusion of NLS-GFP.

ABSTRACT: Since its initial development in 1976, fluorescence recovery after photobleaching (FRAP) has been one of the most popular tools for studying diffusion and protein dynamics in living cells. Its popularity is derived from the widespread availability of confocal microscopes and the relative ease of the experiment and analysis. FRAP, however, is limited in its ability to resolve spatial heterogeneity. Here, we combine selective plane illumination microscopy (SPIM) and FRAP to create SPIM-FRAP, wherein we use a sheet of light to bleach a two-dimensional (2D) plane and subsequently image the recovery of the same image plane. This provides simultaneous quantification of diffusion or protein recovery for every pixel in a given 2D slice, thus moving FRAP measurements beyond these previous limitatio

SUBMITTER: Hobson CM 

PROVIDER: S-EPMC7399492 | biostudies-literature | 2020 Aug

REPOSITORIES: biostudies-literature