( dually and thereby localize its center...)New light microscope sharpens scientists' focus

dually and thereby localize its center, Hess said. When a final image is created that includes the center of each individual molecule, it has a resolution previously only achievable with an electron microscope. Unlike electron microscopy, however, the new technique allows for more flexibility in labeling molecules of interest.

By early 2006, the collaborators were taking strikingly clear images of a number of cellular structures, including actin filaments, focal adhesion proteins, mitochondria, and the Golgi apparatus. "Performance-wise, it's just astounding. And simplicity-wise it's astounding," Betzig said. "It knocks the socks off of standard fluorescence microscopy from a resolution perspective."

Hess added, "This level of detail was previously only obtainable with an electron microscope. However, the contrast in electron microscopy is more indiscriminate, whereas we can limit our contrast to only specific proteins of interest."

Lippincott-Schwartz pointed out that although there are advantages to both imaging techniques, the use of PALM in conjunction electron microscopy is particularly powerful. "A great feature of PALM is that is can readily be used with electron microscopy, which produces a detailed image of very small structures ?but not proteins ?in cells," she said. "By correlating a PALM image showing protein distribution with an electron microscope image showing cell structure of the same sample, it becomes possible to understand how molecules are individually distributed in a cellular structure at the molecular scale. Correlative PALM unites the advantages of light and electron microscopy, producing a revolutionary new approach for looking at the cell in molecular detail."

For now, Hess and Betzig have focused on creating PALM images with a single fluorescent protein, which limits them to mapping a single target protein per sample. "Right now the resolution is wonderful, but it's all single-label stuff. Knowing
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Source:Howard Hughes Medical Institute

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