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*****
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Dear Alan--
On 8/31/2012 7:22 AM, Alan Smith wrote:
> I understand how the illumination psf is formed in STED using a depletion
> beam. However, if the emission is detected is collected in epi-detection,
> why is the resolution not determined by the diffraction limit for the objective.
>
> As an example, if a single molecule was producing fluorescence, the image
> will be an airy disk determined by the NA of the detecting objective and the
> wavelength of light. Despite the fluorescence solely coming from a single
> molecule.
You're right: with both a single molecule and with STED, you see a
diffraction-limited Airy disk on the emission side. However, with STED,
you know to a high degree of precision the position of the excitation
beam eliciting that Airy disk. Thus, as you scan that small excitation
beam over a small structure, you can obtain resolution that's many times
better than confocal. In that way, it's similar to near-field
super-resolution methods.
Hope that helps!
Martin Wessendorf
--
Martin Wessendorf, Ph.D. office: (612) 626-0145
Assoc Prof, Dept Neuroscience lab: (612) 624-2991
University of Minnesota Preferred FAX: (612) 624-8118
6-145 Jackson Hall, 321 Church St. SE Dept Fax: (612) 626-5009
Minneapolis, MN 55455 e-mail: [log in to unmask]
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