Introduction

Spectral imaging with polarity-sensitive fluorescent probes enables the quantification of cell and model membrane physical properties, including local hydration, fluidity, and lateral lipid packing, usually characterized by the generalized polarization (GP) parameter. With the development of commercial microscopes equipped with spectral detectors, spectral imaging has become a convenient and powerful technique for measuring GP and other membrane properties. The existing tools for spectral image processing, however, are insufficient for processing the large data sets afforded by this technological advancement, and are unsuitable for processing images acquired with rapidly internalized fluorescent probes.Here we present a MATLAB spectral imaging toolbox with the aim of overcoming these limitations. In addition to common operations, such as the calculation of distributions of GP values, generation of pseudo-colored GP maps, and spectral analysis, a key highlight of this tool is reliable membrane segmentation for probes that are rapidly internalized. Furthermore, handling for hyperstacks, 3D reconstruction and batch processing facilitates analysis of data sets generated by time series, z-stack, and area scan microscope operations. Finally, the object size distribution is determined, which can provide insight into the mechanisms underlying changes in membrane properties and is desirable for e.g. studies involving model membranes and surfactant coated particles. Analysis is demonstrated for cell membranes, cell-derived vesicles, model membranes, and microbubbles with environmentally-sensitive probes Laurdan, carboxyl-modified Laurdan (C-Laurdan), Di-4-ANEPPDHQ, and Di-4-AN(F)EPPTEA (FE), for quantification of the local lateral density of lipids or lipid packing.The Spectral Imaging Toolbox is a powerful tool for the segmentation and processing of large spectral imaging datasets with a reliable method for membrane segmentation and no ability in programming required. The Spectral Imaging Toolbox can be downloaded from https://uk.mathworks.com/matlabcentral/fileexchange/62617-spectral-imaging-toolbox .

Publications

  1. Spectral imaging toolbox: segmentation, hyperstack reconstruction, and batch processing of spectral images for the determination of cell and model membrane lipid order.
    Cite this
    Aron M, Browning R, Carugo D, Sezgin E, Bernardino de la Serna J, Eggeling C, Stride E, 2017-05-01 - BMC bioinformatics

Credits

  1. Miles Aron
    Developer

    Department of Engineering Science, Institute of Biomedical Engineering

  2. Richard Browning
    Developer

    Department of Engineering Science, Institute of Biomedical Engineering

  3. Dario Carugo
    Developer

    Faculty of Engineering and The Environment, University of Southampton

  4. Erdinc Sezgin
    Developer

    MRC Human Immunology Unit, Weatherall Institute of Molecular Medicine

  5. Jorge Bernardino de la Serna
    Developer

    Research Complex at Harwell, Central Laser Facility

  6. Christian Eggeling
    Developer

    MRC Human Immunology Unit, Weatherall Institute of Molecular Medicine

  7. Eleanor Stride
    Investigator

    Department of Engineering Science, Institute of Biomedical Engineering

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Summary
AccessionBT001763
Tool TypeApplication
Category
PlatformsWindows
Technologies
User InterfaceTerminal Command Line
Download Count0
Submitted ByEleanor Stride