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Originally published in Science Express on 10 August 2006
Science 15 September 2006:
Vol. 313. no. 5793, pp. 1642 - 1645
DOI: 10.1126/science.1127344
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Reports

Imaging Intracellular Fluorescent Proteins at Nanometer Resolution

Eric Betzig,1,2*{dagger} George H. Patterson,3 Rachid Sougrat,3 O. Wolf Lindwasser,3 Scott Olenych,4 Juan S. Bonifacino,3 Michael W. Davidson,4 Jennifer Lippincott-Schwartz,3 Harald F. Hess5*

We introduce a method for optically imaging intracellular proteins at nanometer spatial resolution. Numerous sparse subsets of photoactivatable fluorescent protein molecules were activated, localized (to ~2 to 25 nanometers), and then bleached. The aggregate position information from all subsets was then assembled into a superresolution image. We used this method—termed photoactivated localization microscopy—to image specific target proteins in thin sections of lysosomes and mitochondria; in fixed whole cells, we imaged vinculin at focal adhesions, actin within a lamellipodium, and the distribution of the retroviral protein Gag at the plasma membrane.

1 Howard Hughes Medical Institute, Janelia Farm Research Campus, Ashburn, VA 20147, USA.
2 New Millennium Research, LLC, Okemos, MI 48864, USA.
3 Cell Biology and Metabolism Branch, National Institute of Child Health and Human Development (NICHD), Bethesda, MD 20892, USA.
4 National High Magnetic Field Laboratory, Florida State University, Tallahassee, FL 32310, USA.
5 NuQuest Research, LLC, La Jolla, CA 92037, USA.

* These authors contributed equally to this work.

{dagger} To whom correspondence should be addressed. E-mail: betzige{at}janelia.hhmi.org.

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Science. ISSN 0036-8075 (print), 1095-9203 (online)