Re: Estimating field of view in an LSM
Posted by
Julio Vazquez on
URL: http://confocal-microscopy-list.275.s1.nabble.com/Estimating-field-of-view-in-an-LSM-tp590288p590296.html
Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
-
Hi Craig,
I'm venturing into long-faded and possibly confused memories here, but I'm inclined to think that you can determine this from geometrical optics. Basically, a parallel bundle of rays that enters the lens at a perpendicular (normal) angle will be focused to the focal spot (center of the field of view), while a parallel bundle of rays that enters the lens at an angle alpha along one of the axes (alpha being equal to the half-cone of acceptance, basically the angle used to measure NA) will be focused at a distance from the center of the field such that d/f = tan(alpha) . You can verify this by a simple ray diagram. For any arbitrary angle you would have to consider the deviation along both axes to know exactly where the beam will be focused. I don't know if there are additional subtleties when more realistic "thick lenses", "compound lenses" or infinity lenses are involved...
Actually, I just consulted the Confocal Handbook, 3rd Edition, to make sure I was not talking nonsense, and Chapter 9 contains a diagram and gives some formulas that seem to confirm the above statement... You may want to check it out, specifically pp 208-209.
I would assume that in practice the actual field of view will be limited by the dimensions of the laser beam and objective, and the size of the scanning mirror, since at some point, you can't increase the angle without losing power, getting some "vignetting-like" effect, or getting geometrical distortions...
Just for reference, on our Zeiss LSM 510, the field dimensions with a 20x lens at zoom 1 are 460.7 x 460.7 microns, and they would be half that much with a 40x or with a 20x at zoom 2 (independently of NA). On our Leica SP, the values are 500x500 microns for 20x at zoom 1, etc...
--
Julio Vazquez,
Fred Hutchinson Cancer Research Center
1100 Fairview Ave. N., mailstop DE-512
Seattle, WA 98109-1024
On Sep 21, 2007, at 1:29 PM, Craig Brideau wrote:
Search the CONFOCAL archive at
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
Search the CONFOCAL archive at
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.
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
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
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