Bayesian localization microscopy reveals nanoscale podosome dynamics

A localization microscopy analysis method that is described
able to extract results in live cells using standard fluorescent
proteins and xenon arc lamp illumination. The Bayesian analysis
of the blinking and bleaching (3B analysis) method models the
entire dataset simultaneously as being generated by a number
of fluorophores that may or may not be emitting light at any
given time. The resulting technique allows many overlapping
fluorophores in each frame and unifies the analysis of the
localization from blinking and bleaching events. By modeling
the entire dataset, we were able to use each reappearance
of a fluorophore to improve the localization accuracy. The
high performance of this technique allowed us to reveal the
nanoscale dynamics of podosome formation and dissociation
throughout an entire cell with a resolution of 50 nm on a 4-s
timescale.

Cox S et al, Nature Methods,

PUBLISHED ONLINE 4 DECEMBER 2011; DOI:10.1038/NMETH.1812

 

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