Re: Estimating field of view in an LSM

Posted by Guy Cox on
URL: http://confocal-microscopy-list.275.s1.nabble.com/Estimating-field-of-view-in-an-LSM-tp590288p590290.html

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I'm not sure I see the problem.  Surely
you can find or borrow one lens at least -
the rest is just scaling.  Rather than buy
an expensive test sample I'd suggest
bleaching an area on a disposable sample
and measuring the bleached area.  Or for
a low-power objective mark some lines with
a fine black pen on one of the Chroma
fluorescent plastic slides.  

If you really want to do it by ray-tracing
I suggest not doing it through the objective
(since, as you say, it's not quite clear where
the bfp of the objective is) but transferring
your angles to displacements at the primary image
plane - you can rely on the objective to do a
faithful demagnification of the image plane to
the sample - of course this does asssume you are
forming your image at the correct plane.  

                                   Guy



Optical Imaging Techniques in Cell Biology
by Guy Cox    CRC Press / Taylor & Francis
    http://www.guycox.com/optical.htm
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Associate Professor Guy Cox, MA, DPhil(Oxon)
Electron Microscope Unit, Madsen Building F09,
University of Sydney, NSW 2006
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-----Original Message-----
From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Craig Brideau
Sent: Saturday, 22 September 2007 6:30 AM
To: [hidden email]
Subject: Re: Estimating field of view in an LSM

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Yeah, I want to be able to predict the field ahead of time BEFORE I buy an objective.  I'll have to keep looking in to the theory.  If it was a simple lens I could just use some ray trace equations, but microscope objectives are very much 'black box' devices due to their complexity, and proprietary design.  Still, good advice for characterizing my existing lenses.

Thanks,

Craig

On 9/21/07, Steffen Dietzel <[hidden email]> wrote:

> Search the CONFOCAL archive at
> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>
> Craig,
>
> for the theory I don't know, but to actually meassure the field of
> view (or the size of your pixels) you can use a 1951 USAF Resolution
> Test Chart. See e.g.
> http://en.wikipedia.org/wiki/1951_USAF_Resolution_Test_Chart
>
> There are several suppliers you can google for. I got mine from Edmund
> Optics. It has a mirror coating on the top surface so that it can be
> used in reflection mode.
>
> Steffen
>
>
>
> At 22:16 20.09.2007, you wrote:
> >Search the CONFOCAL archive at
> >http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
> >
> >Just to clarify, I am doing multiphoton with a single PMT, so I'm
> >trying to measure the field that the laser is focused over at the
> >focal plane of the objective for a given input beam deflection.
> >
> >Craig
> >
> >On 9/20/07, Craig Brideau <[hidden email]> wrote:
> > > I was wondering how I estimate the field of view for a given lens
> > > in a laser scanning microscope?  I know it will depend on the
> > > entrance angle of the beam into the back aperture of the lens, but
> > > I don't know how to translate this into lateral focal point
> > > displacement for a given working distance/NA/magnification of
> > > lens.  Can anyone point me towards some good references?
> > >
> > > Thanks,
> > >
> > > Craig
> > >
>

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