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Published Online August 10, 2006
Science DOI: 10.1126/science.1127344

Reports

Submitted on March 13, 2006
Accepted on August 2, 2006

Imaging Intracellular Fluorescent Proteins at Nanometer Resolution

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

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

* To whom correspondence should be addressed.
Eric Betzig , E-mail: betzige{at}hhmi.org

A method for optically imaging intracellular proteins at nanometer spatial resolution is introduced. Numerous sparse subsets of photoactivatable fluorescent protein molecules are activated, localized (to ~2-25 nm), and then bleached. The aggregate position information from all subsets is then assembled into a superresolution image. The method, termed photoactivated localization microscopy (PALM), is demonstrated in thin sections by imaging specific target proteins in lysosomes and mitochondria, and in fixed, whole cells by imaging vinculin at focal adhesions, actin within a lamellipodium, and the distribution of the retroviral protein Gag at the plasma membrane.


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