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Int. J. Mol. Sci. 2013, 14(8), 16485-16514; doi:10.3390/ijms140816485

Correlation of Dual Colour Single Particle Trajectories for Improved Detection and Analysis of Interactions in Living Cells

1
Laboratory of General Biochemistry and Physical Pharmacy, Ghent University, Harelbekestraat 72, B-9000 Gent, Belgium
2
Center for Nano- and Biophotonics, Ghent University, B-9000 Gent, Belgium
3
Liquid Crystals and Photonics Group, Ghent University, Sint-Pietersnieuwstraat 41, B-9000 Gent, Belgium
*
Author to whom correspondence should be addressed.
Received: 12 June 2013 / Revised: 18 July 2013 / Accepted: 29 July 2013 / Published: 8 August 2013
(This article belongs to the Special Issue Frontiers of Micro-Spectroscopy in Biological Applications)
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Abstract

Interactions between objects inside living cells are often investigated by looking for colocalization between fluorescence microscopy images that are recorded in separate colours corresponding to the fluorescent label of each object. The fundamental limitation of this approach in the case of dynamic objects is that coincidental colocalization cannot be distinguished from true interaction. Instead, correlation between motion trajectories obtained by dual colour single particle tracking provides a much stronger indication of interaction. However, frequently occurring phenomena in living cells, such as immobile phases or transient interactions, can limit the correlation to small parts of the trajectories. The method presented here, developed for the detection of interaction, is based on the correlation inside a window that is scanned along the trajectories, covering different subsets of the positions. This scanning window method was validated by simulations and, as an experimental proof of concept, it was applied to the investigation of the intracellular trafficking of polymeric gene complexes by endosomes in living retinal pigment epithelium cells, which is of interest to ocular gene therapy. View Full-Text
Keywords: interaction; fluorescence microscopy; single particle tracking; colocalization; correlation; endosomal escape; diffusion interaction; fluorescence microscopy; single particle tracking; colocalization; correlation; endosomal escape; diffusion
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This is an open access article distributed under the Creative Commons Attribution License (CC BY 3.0).

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MDPI and ACS Style

Deschout, H.; Martens, T.; Vercauteren, D.; Remaut, K.; Demeester, J.; De Smedt, S.C.; Neyts, K.; Braeckmans, K. Correlation of Dual Colour Single Particle Trajectories for Improved Detection and Analysis of Interactions in Living Cells. Int. J. Mol. Sci. 2013, 14, 16485-16514.

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