Recent experiments and simulations are starting to answer some fundamental questions about how life came to be.
Image credit: Shutterstock/Radoslaw Lecyk.
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Contributed by Jennifer Lippincott-Schwartz, December 25, 2008 (received for review November 19, 2008)
Abstract
Understanding molecular-scale architecture of cells requires determination of 3D locations of specific proteins with accuracy matching their nanometer-length scale. Existing electron and light microscopy techniques are limited either in molecular specificity or resolution. Here, we introduce interferometric photoactivated localization microscopy (iPALM), the combination of photoactivated localization microscopy with single-photon, simultaneous multiphase interferometry that provides sub-20-nm 3D protein localization with optimal molecular specificity. We demonstrate measurement of the 25-nm microtubule diameter, resolve the dorsal and ventral plasma membranes, and visualize the arrangement of integrin receptors within endoplasmic reticulum and adhesion complexes, 3D protein organization previously resolved only by electron microscopy. iPALM thus closes the gap between electron tomography and light microscopy, enabling both molecular specification and resolution of cellular nanoarchitecture.
- fluorescence microscopy
- interferometry
- PALM
- photoactivated localization microscopy
- single molecule imaging
Footnotes
- 1To whom correspondence may be addressed. E-mail: hessh{at}janelia.hhmi.org or jlippin{at}helix.nih.gov
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Author contributions: G.S., J.A.G., C.G.G., J.L.-S., J.M.G., S.M., R.S., C.M.W., P.K., and H.F.H. designed research; G.S., J.A.G., C.G.G., J.M.G., S.M., R.S., P.K., R.D.F., and H.F.H. performed research; G.S., J.A.G., C.G.G., J.M.G., S.M., P.K., M.W.D., and H.F.H. contributed new reagents/analytic tools; G.S., J.A.G., C.G.G., J.M.G., S.M., P.K., and H.F.H. analyzed data; and G.S., J.A.G., C.G.G., J.L.-S., J.M.G., S.M., C.M.W., P.K., M.W.D., R.D.F., and H.F.H. wrote the paper.
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The authors declare no conflict of interest.
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This article contains supporting information online at www.pnas.org/cgi/content/full/0813131106/DCSupplemental.
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Freely available online through the PNAS open access option.
Interferometric fluorescent super-resolution microscopy resolves 3D cellular ultrastructure
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