Posted by
Kevin Ryan on
URL: http://confocal-microscopy-list.275.s1.nabble.com/Question-about-deconvolution-tp7579203p7579225.html
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It's important to note that there are a few different issues here.
The _theoretic_ resolution is driven by the highest observed spatial frequencies, the sharpest edges, that make it through the observing system. This is set by the aperture and wavelength of the observation, an information limit.
The _effective_ resolution, the ability to separate nearby objects, is also dependent on the _strength_ of those frequencies. In a single in focus XY plane (for example), high frequencies drop in strength from 100% at the average value, to zero at the high frequency aperture limit, the sharpest edges. That fall-off blurs the observations considerably.
Deconvolution restores higher frequency strengths (in frequency terms, changing a tapered pyramid centered at zero to a near-box), reducing the full-width half max (FWHM) of observed structures accordingly. You have more edge information, which means sharper edges. Add to that (in widefield) the removal of uncorrelated out of focus illumination, and (particularly in confocal) reduction of noise that's unsupported by a PSF, and you will obtain a considerable improvement in _effective_ resolution, the ability to separate nearby structures.
Deconvolution won't provide superresolution (frequencies past the aperture limit aren't represented in the observation, and in a noise-limited real world situation any estimate would be unstable), but the effective observed deconvolution resolution improvement, separability, in XYZ is ~2-2.5x over the original data.
See Kano 1996 (
http://www3.mpibpc.mpg.de/groups/hell/publications/pdf/Bioimaging_4_187-197.pdf - thank you for that reference, Hans) for a discussion of FWHM improvements seen in maximum-likelyhood restoration.
Kevin Ryan
Media Cybernetics, Inc.
-----Original Message-----
From: Confocal Microscopy List [mailto:
[hidden email]] On Behalf Of Christophe Leterrier
Sent: Tuesday, October 23, 2012 12:28 PM
To:
[hidden email]
Subject: Question about deconvolution
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Hi folks,
I have a long-standing question regarding deconvolution (as processing widefield or confocal images to reassign light from where it originated using a PSF).
Is there a theoretical limit to the resolution one could obtain using deconvolution? Is is theoretically possible to "break" the diffraction limit with deconvolution? That is, to get under the classical 200x200x600nm spot? I think it is not the case, but then why would you deconvolve widefield or confocal images? What do you gain by doing so on a system that is reasonably close to its theoretical capabilities in terms of optical performances?
Thanks for your help,
Christophe
--
Christophe Leterrier
Researcher
Axonal Domains Architecture Team
CRN2M CNRS UMR 7286
Aix Marseille University, France
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