Journal: Biology Open
Article Title: Computational tools for quantifying actin filament numbers, lengths, and bundling
doi: 10.1242/bio.060267
Figure Lengend Snippet: Kinetic measurements of filament bundling. Filaments were assembled from 2 µM actin monomers, labeled with FITC-phalloidin, and visualized by TIRF microscopy following the addition of 1.4 µM fascin. (A) Time series of micrographs depicting the progress of filament bundling. The same micrographs are also shown in Figure 4 to demonstrate the effects of image segmentation on the resolution of filaments and bundles. (B) Micrograph collected at time=0 s, before bundling has occurred. Following background subtraction, noise filtering, thresholding and skeletonization, the image has been segmented into a grid. The numbers correspond to the threshold fluorescence value (reported in arbitrary units of fluorescence intensity) above which pixels are considered bundled within each segment of the grid. (C) Automated detection of stretches of bundled (yellow) and single (magenta) filaments in each of the micrographs shown in panel A. (D) The fraction of the filamentous actin that is bundled over time for the reaction shown in panel A.
Article Snippet: A thresholding algorithm (implemented using MATLAB's ‘graythresh’ and ‘imbinarize’ functions ( https://www.mathworks.com/help/images/ref/graythresh.html ; https://www.mathworks.com/help/images/ref/imbinarize.html ) is then applied to convert detected objects from a grayscale into binary images ( ).
Techniques: Labeling, Microscopy, Fluorescence