114 research outputs found
A photoactivatable marker protein for pulse-chase imaging with superresolution
No full textIrisFP is a photoactivatable fluorescent protein that combines irreversible photoconversion from a green- to a red-emitting form with reversible photoswitching between a fluorescent and a nonfluorescent state in both forms. Here we introduce a monomeric variant, mIrisFP, and demonstrate how its multiple photoactivation modes can be used for pulse-chase experiments combined with subdiffraction-resolution imaging in living cells by using dual-color photoactivation localization microscopy (PALM)
Visualization of barriers and obstacles to molecular diffusion in live cells by spatial pair-cross-correlation in two dimensions
Full text linkDespite recent advances in optical super-resolution, we lack a method that can visualize the path followed by diffusing molecules in the cytoplasm or in the nucleus of cells. Fluorescence correlation spectroscopy (FCS) provides molecular dynamics at the single molecule level by averaging the behavior of many molecules over time at a single spot, thus achieving very good statistics but at only one point in the cell. Earlier image-based methods including raster-scan and spatiotemporal image correlation need spatial averaging over relatively large areas, thus compromising spatial resolution. Here, we use spatial pair-crosscorrelation in two dimensions (2D-pCF) to obtain relatively high resolution images of molecular diffusion dynamics and transport in live cells. The 2D-pCF method measures the time for a particle to go from one location to another by cross-correlating the intensity fluctuations at specific points in an image. Hence, a visual map of the average path followed by molecules is created
Phasor-based hyperspectral snapshot microscopy allows fast imaging of live, three-dimensional tissues for biomedical applications
Full text linkHedde et al. demonstrate the use of ultrafast phasor-based hyperspectral snapshot microscopy for biomedical imaging. This technique can improve imaging speed by 10-100 fold and enables 3D hyperspectral imaging of live tissues without using expensive and specialized hyperspectral cameras
Diffusion Mapping in Living Cells using Camera-Based Correlation Spectroscopy and Phasor Analysis
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Selective plane illumination microscopy with a light sheet of uniform thickness formed by an electrically tunable lens
Full text linkLight sheet microscopy is a powerful technique for rapid, three-dimensional fluorescence imaging of large specimen such as drosophila and zebrafish embryos. Yet, beam divergence results in a loss of axial resolution at the periphery of the light sheet. Here, we demonstrate how an electrically tunable lens can be utilized to maintain the minimal, diffraction-limited thickness of the light sheet over a wide field of view (>600 µm) at high frame rates (40 fps). This mode of operation is necessary for the application of fluorescence fluctuation spectroscopy in images. Microsc. Res. Tech. 81:924-928, 2018. © 2016 Wiley Periodicals, Inc
Selective plane illumination microscopy with a light sheet of uniform thickness formed by an electrically tunable lens
No full textDiffusion Mapping in Living Cells using Camera-Based Correlation Spectroscopy and Phasor Analysis
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