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Article
Nature Methods - 5, 417 - 423 (2008)
Published online: 13 April 2008; | doi:10.1038/nmeth.1202

Live-cell photoactivated localization microscopy of nanoscale adhesion dynamics

Hari Shroff1, 4, Catherine G Galbraith2, 4, James A Galbraith3, 4 & Eric Betzig1

1  Janelia Farm Research Campus, Howard Hughes Medical Institute, 19700 Helix Drive, Ashburn, Virginia 20147, USA.

2  National Institute of Dental and Craniofacial Research, National Institutes of Health, 30 Convent Drive, Bethesda, Maryland 20892, USA.

3  National Institute of Neurological Disorders and Stroke, National Institutes of Health, 49 Convent Drive, Bethesda, Maryland 20892, USA.

4  These authors contributed equally to this work.

Correspondence should be addressed to Hari Shroff shroffh@janelia.hhmi.org

We demonstrate live-cell super-resolution imaging using photoactivated localization microscopy (PALM). The use of photon-tolerant cell lines in combination with the high resolution and molecular sensitivity of PALM permitted us to investigate the nanoscale dynamics within individual adhesion complexes (ACs) in living cells under physiological conditions for as long as 25 min, with half of the time spent collecting the PALM images at spatial resolutions down to approx60 nm and frame rates as short as 25 s. We visualized the formation of ACs and measured the fractional gain and loss of individual paxillin molecules as each AC evolved. By allowing observation of a wide variety of nanoscale dynamics, live-cell PALM provides insights into molecular assembly during the initiation, maturation and dissolution of cellular processes.

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Nature Methods
ISSN: 1548-7091
EISSN: 1548-7105
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