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Hi Andrew
All rays whose effective NA >1 get reflected back by
the system you
describe. There are more high NA rays
than low NA rays (due to the area
of an annulus
increasing with it's radius : pi(2r dr). As I see it, one
can't measure the true throughput with axial rays as the marginal rays
encouner very different glass thicknesses and there may
be internal
field stops (or even phase rings!). The
measurement of throughput is
certainly not going to be
easy. I don't see how this is related to the
sine
condition, which I believe simply defines a geometric constraint
for an undistorted image viz:
http://en.wikipedia.org/wiki/Abbe_sine_condition
Cheers Mark
Andrew Resnick wrote:
> Search the
CONFOCAL archive at
> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>
> Mark,
>
> That's a good question. We
attached a coverslip with a small drop of
> oil to
the objective as a way to deal with immersion objectives. I
> don't know what it means that the "sample" is in
air; if some light is
> then totally internally
reflected back into the coverslip, which could
> be a
spectral issue due to dispersion.
>
> Using two matched objectives is probably a better method, but I
> imagine considerably more difficult due to
alignment constraints and
> obtaining matched
objectives. The objectives we had obeyed the 'sine
> condition', which I believe means that there are fewer high-NA rays
> than low NA rays- this would help mitigate our
experimental error as
> well.
>
>
> At 04:19
PM 9/2/2007, you wrote:
>> Search the CONFOCAL
archive at
>> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>>
>> Hi Andrew
>>
>> While I can see that
method would work for lenses designed to work in
>> air, wouldn't total internal reflection be a problem for lenses
with
>> an NA over 1.0? The only way I can think
of doing it is to use two
>> objectives of the
same type looking at each other at a common focal
>> point with the right immersion medium between them. Then the
overall
>> transmission would be half that of a
single lens...
>>
>>
Cheers Mark
>>
>>
Andrew Resnick wrote:
>>> Search the CONFOCAL
archive at
>>> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>>>
>>> We've done
similar measurements, it's not too difficult.
>>>
>>> The main trick is
handling the large NA lenses. Our setup was light
>>> source -> objective -> integrating sphere ->
spectrometer. After
>>> normalizing to
the source, we obtained really good data. It's
>>> pretty easy, actually.
>>>
>>> Andy
>>>
>>>
>>> At 08:45 AM 8/31/2007, you wrote:
>>>> Search the CONFOCAL archive at
>>>> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>>>>
>>>> Dear
all,
>>>>
>>>> one of the main differences of objectives is their
transmission
>>>> efficiency at certain
wavelengths. One way to compare this is the
>>>> trial-and-error method, however, this is not straight
forward.
>>>>
>>>> My idea is to use a combination of spectrophotometer and
a lamp
>>>> with a more or less even spectra
(i.e. Xenon) on some kind of
>>>> optical
bench. This would make the setup independent from the
>>>> manufacturer. Beside the distance between the light
source and the
>>>> detector, there are
obviously more things to consider: different
>>>> diameter of the back focal plane, different focal
lenghts...
>>>>
>>>> I would like to hear about your opinion about how to
measure
>>>> objective transmission. Have
you ever done this in your lab? Did
>>>> you
find a setup that worked for you?
>>>>
>>>> cheers,
>>>> Michael
>>>
>>> Andrew Resnick, Ph.
D.
>>> Instructor
>>> Department of Physiology and Biophysics
>>> Case Western Reserve University
>>> 216-368-6899 (V)
>>>
216-368-4223 (F)
>>
>> Andrew Resnick, Ph. D.
>>
Instructor
>> Department of Physiology and
Biophysics
>> Case Western Reserve
University
>> 216-368-6899 (V)
>> 216-368-4223 (F)
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