Posted by
Guy Cox-2 on
URL: http://confocal-microscopy-list.275.s1.nabble.com/why-does-high-NA-excitation-illumination-give-better-resolution-in-fluorescence-microscopy-tp7581519p7581527.html
*****
To join, leave or search the confocal microscopy listserv, go to:
http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy*****
I think SA would negate any resolution improvement. But what many people have suggested is that these people are getting 'accidental TIRF' - in other words, TIRF-enhanced fluorescence of structures very close to the coverslip. I don't know of any serious studies investigating this.
Guy
-----Original Message-----
From: Confocal Microscopy List [mailto:
[hidden email]] On Behalf Of Steffen Dietzel
Sent: Wednesday, 5 February 2014 4:56 AM
To:
[hidden email]
Subject: Re: why does high NA excitation illumination give better resolution in fluorescence microscopy?
*****
To join, leave or search the confocal microscopy listserv, go to:
http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy*****
Am 03.02.2014 18:39, schrieb Reto Fiolka:
> I have seen many papers where people employed high NA oil objectives for
> confocal imaging in biological samples. Besides not being indexed matched, they
> will not provide more resolution, as any illumination above the critical angle is
> lost in the near field (critical angle for n=1.33 is reached at NA 1.33).
Well, NA 1.33 does sound like better resolution than the NA 1.2 of the
best Water-Coverslip-Objectives, does it not?
And just for the fun of it, if cells grow directly on the coverslip, n
is somewhat larger than 1.33 since you stay inside the cell for the
important parts of the image. (not that I would expect this to make a
big difference though)
Steffen
--
------------------------------------------------------------
Steffen Dietzel, PD Dr. rer. nat
Ludwig-Maximilians-Universität München
Walter-Brendel-Zentrum für experimentelle Medizin (WBex)
Head of light microscopy
Mail room:
Marchioninistr. 15, D-81377 München
Building location:
Marchioninistr. 27, München-Großhadern