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Segmentation and quantification of subcellular structures in fluorescence microscopy images using Squassh.

MPS-Authors

Rizk,  Aurelien
Max Planck Society;

Paul,  Gregory
Max Planck Society;

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Incardona,  Pietro
Max Planck Institute of Molecular Cell Biology and Genetics, Max Planck Society;

Bugarski,  Milica
Max Planck Society;

Mansouri,  Maysam
Max Planck Society;

Niemann,  Axel
Max Planck Society;

Ziegler,  Urs
Max Planck Society;

Berger,  Philipp
Max Planck Society;

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Sbalzarini,  Ivo F.
Max Planck Institute of Molecular Cell Biology and Genetics, Max Planck Society;

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Citation

Rizk, A., Paul, G., Incardona, P., Bugarski, M., Mansouri, M., Niemann, A., et al. (2014). Segmentation and quantification of subcellular structures in fluorescence microscopy images using Squassh. Nature Protocols, 9(3), 586-596.


Cite as: http://hdl.handle.net/21.11116/0000-0001-0608-0
Abstract
Detection and quantification of fluorescently labeled molecules in subcellular compartments is a key step in the analysis of many cell biological processes. Pixel-wise colocalization analyses, however, are not always suitable, because they do not provide object-specific information, and they are vulnerable to noise and background fluorescence. Here we present a versatile protocol for a method named 'Squassh' (segmentation and quantification of subcellular shapes), which is used for detecting, delineating and quantifying subcellular structures in fluorescence microscopy images. The workflow is implemented in freely available, user-friendly software. It works on both 2D and 3D images, accounts for the microscope optics and for uneven image background, computes cell masks and provides subpixel accuracy. The Squassh software enables both colocalization and shape analyses. The protocol can be applied in batch, on desktop computers or computer clusters, and it usually requires <1 min and <5 min for 2D and 3D images, respectively. Basic computer-user skills and some experience with fluorescence microscopy are recommended to successfully use the protocol.