Re: [External] Re: Numerical aperture and spatial resolution

Posted by JOEL B. SHEFFIELD on
URL: http://confocal-microscopy-list.275.s1.nabble.com/Numerical-aperture-and-spatial-resolution-tp7591293p7591301.html

*****
To join, leave or search the confocal microscopy listserv, go to:
http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
Post images on http://www.imgur.com and include the link in your posting.
*****

About 20 years ago, Jim Pawley and Guy Cox developed a wonderful simulation
of the original Abbe experiment along with a clear analysis of the optics
involved. It focuses (pardon the pun) on the diffraction component rather
than the wavelength.   You can "mount" a slide with a grating as a
specimen, and insert a slider at the back focal plane to block orders lines.
If you can still get it, I suppose Guy Cox would be the resource.
Alternatively, I have the files if someone is interested.

I should say that in my microscopy course, I have the students use several
samples with fine repeats, a stage micrometer, a hemocytometer, some trick
"rainbow" glasses, and I have them look at the back focal plane to see
what's going on.

Joel


Joel B. Sheffield, Ph.D
Department of Biology
Temple University
Philadelphia, PA 19122
Voice: 215 204 8839
e-mail: [hidden email]
Office:  Biolife 311
URL:  *https://bio.cst.temple.edu/~jbs/ <https://bio.cst.temple.edu/~jbs/>
<http://tinyurl.com/khbouft>*


On Sat, Sep 19, 2020 at 3:22 PM F Javier Diez Guerra <[hidden email]>
wrote:

> *****
> To join, leave or search the confocal microscopy listserv, go to:
> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> Post images on http://www.imgur.com and include the link in your posting.
> *****
>
> Hi,
>
> Thanks to all contributors for the excellent suggestions.
>
> Unfortunately, at the present times, at least here in Madrid, most if
> not all university lectures are Teams sessions. So on-site demos are not
> a choice.
>
> I also align with Zdenee's view. "Why does higher NA mean higher
> resolution?" is a question with no trivial answer for biologists. The
> demo of looking the diffraction orders at the BFP with a Bertrand lens,
> using objectives with different NA and gratings of different densities,
> worked best for me when I first learned about spatial resolution. Still,
> for a biology undergraduate is difficult to comprehend intuitively why
> capturing more diffraction orders means higher resolution.
>
> Javier
>
>
> El 19/09/2020 a las 20:27, Zdenek Svindrych escribió:
> > *****
> > To join, leave or search the confocal microscopy listserv, go to:
> > http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> > Post images on http://www.imgur.com and include the link in your
> posting.
> > *****
> >
> > Hi George and Javier,
> >
> > regarding aperture and resolution of DSLR (actually mirror-less) I put
> some
> > photos together here:
> > https://drive.google.com/file/d/1vDYqDBPgapZYad3JahRm8R5W5Bkwu4T6
> > But, it will only work with some lenses (well, with most, but not in the
> > full F-stop range), and only with some cameras (more pixels is better
> here,
> > kind of an exception :-).
> >
> > Davide, the Evenett's video is great! Many demonstrations were quite
> > surprising, and I have nothing to add to it (maybe a note, that the phase
> > contrast method in all its simplicity still yielded Zernike a Nobel
> prize!).
> > But even though one thought follows from another nicely, at the end if
> you
> > ask your students "So why does higher NA mean higher resolution?" they'll
> > say "Well, uhm..." It's not very likely they'll gain a deeper
> understanding
> > of how all these concepts fit together.
> >
> > Good luck!
> >
> > zdenee
> >
> > On Sat, Sep 19, 2020 at 12:25 PM George McNamara <
> [hidden email]>
> > wrote:
> >
> >> *****
> >> To join, leave or search the confocal microscopy listserv, go to:
> >> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> >> Post images on http://www.imgur.com and include the link in your
> posting.
> >> *****
> >>
> >> Hi Javier,
> >>
> >> I suggest you ask the class if they have any interest in taking photos
> >> with their smartphone or DSLR camera, and if they do, bring in a DSLR
> >> and tripod and computer/projector (if the room does not have that
> >> already), explain f/stop and NA are inverse of each other, and
> >> demonstrate NA (resolution, intensity, depth of field) on the class by
> >> focusing on faces in the middle row. Wavelength: yes, you could buy some
> >> filters for the camera lens (or deal with separating channels in
> >> Photoshop, fiji ImageJ, etc), but probably better done on a microscope.
> >>
> >> George
> >>
> >> p.s. this post was inspired by a conversation yesterday -- Friday happy
> >> hour -- with a colleague, Prof. Jim Potter, who told me about a
> >> conversation he had with someone with a very expensive digital camera
> >> hobby, who only used their cameras set to auto. Jim explained f/stop,
> >> ISO and more (framing scene etc). My thanks to Jim for good story with
> >> nice timing and especially foraging for the beverages.
> >>
> >> A fun (in theory, but probably not practical or wise) wavelength test
> >> would be to use bring in blue (~400 nm) and NIR (~800nm would be nice,
> >> values chosen to be 2 fold, not necessarily practical) and a smoke
> >> machine (and some 400nm and NIR friendly lighting in case room lights
> >> too dim at those wavelength), and demonstration resolution vs ability to
> >> see through the smoke (Mie scattering etc). If you do this, and the
> >> smoke alarm turns on, your school may not be happy with you (though all
> >> the students may be happy to escape class early).
> >>
> >> Of course if they vote no to learning about f/stop and NA, you can ask
> >> them how they expect to get to a million TikTok followers each without
> >> knowing how to take good quality videos. Come ot think of it, if they do
> >> vote yes, and you put your demo on your TikTok feed and make a lot of
> >> money, you can send me the URL and a thank you check.
> >>
> >> On 9/19/2020 5:38 AM, F Javier Diez Guerra wrote:
> >>> *****
> >>> To join, leave or search the confocal microscopy listserv, go to:
> >>> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> >>> Post images on http://www.imgur.com and include the link in your
> >> posting.
> >>> *****
> >>>
> >>> Hello,
> >>>
> >>> I wonder if anybody in the list could help.
> >>>
> >>> I want to convey to biology undergraduate students (very allergic to
> >>> physics and mathematics) the understanding of the relationship between
> >>> numerical aperture and spatial resolution.
> >>>
> >>> I have already given them links to the different microscopy primer
> >>> sites. They find difficult to understand why the airy disk is
> >>> generated in the image plane, how the diffraction orders affect
> >>> resolution and why increasing NA reduces the image spot.
> >>>
> >>> Could anybody share a basic and intuitive infographic slide, animation
> >>> or any other resource that could help to ease comprehension in this
> >>> context?
> >>>
> >>> Thanks
> >>>
> >>> Javier
> >>>
> >>>
> >
>