http://confocal-microscopy-list.275.s1.nabble.com/DIC-condensers-tp7583008p7583018.html
components. You do need a high NA objective lens and digital image
processing. See
p.s. see
> *****
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>
> Dear Jeff,
>
> Guy has already sent an answer and provided good information for you.
>
> I would like to send some additional information:
>
> In case of transmitted light wide field microscopy, as a rule of thumb
> the (lateral!) resolution is
>
> d = (1.28 * lambda) / (NAobj + NAcond)
>
> This formula is quite useful, and shows pretty well, to which degree,
> indeed, resolution is dependent on the numerical aperture of the
> condenser. "Of course", it is never a 100% correct description of
> reality, since you will never be able to attain illumination by "just
> one" wavelength; in the very best case you might get a Lorentzian
> profile of a good single mode laserbeam.
>
> Also, in DIC microscopy, when, as Guy already has written, close to
> perfect Koehler illumnation is a sine qua non for a "good image" -
> whatever THIS would be, :-), another issue is that you can trade
> contrast for resolution. I.e.: By closing the condenser aperture
> diaphragm - often referred to as the "A diaphragm" as opposed to the
> "F diaphragm", which is the Field Diaphragm - you will increase
> contrast while at the same time reduce resolution. In "manuals for
> microscopists" of the sixties and seventies as published in Germany,
> one will sometimes be instructed to do the following: "First, attain
> perfect Koehler illumation, then lower the condenser somewhat to
> attain better contrast." Yes, it works, and it is a kind of rather
> un-controlled oblique illumination, which one fakes in that way. But:
> As a real physicist, you would, for ideological reasons, never do
> this. If you are a pragmatic life scientist: Well, you have a better
> and easier life, anyway.
>
> Also note: Make sure that your light source and your polarizer and
> analyzer are useful in the same wavelength band! At least on older DIC
> systems, polarizers and analyzers will do a good job in the visible
> wavelength range, while they will act as a mere 80% Transmission ND
> filter in IR and do hardly polarize! For IR, you might need separate
> and special polarizers (and analyzers, of course, which are nothing
> else than polarizers in another functional position).
>
> Best wishes and have a good 1st of advent,
>
> Johannes
>
> On 2014-11-28 21:21, Jeff Spector wrote:
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>>
http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy>> Post images on
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>> posting.
>> *****
>>
>> Greetings,
>> This isn't exactly a "confocal" questions but I know a lot of
>> "micoscopy
>> gurus" live on this list so I thought it a good place to ask this. I
>> have a
>> colleague who is trying to image individual (i.e. small and diffraction
>> limited) microtubules in a flow chamber by using DIC. They are currently
>> using a 100x 1.45 oil Objective, but only a .52 LWD condenser. They were
>> using a solid state light source but couldn't get good image so we
>> switched
>> to a lamp for illumination and the images are much better, and we can
>> now
>> see the microtbules but there still isn't a lot of contrast. My
>> question
>> is, is it worth it to go to a high NA (perhaps oil immersion) condenser,
>> and can anyone think of why the lamp would give a better DIC image
>> than a
>> solid state light source?
>> thanks in advance for the help...
>> -Jeff
>
L.J.N. Cooper Lab
University of Texas M.D. Anderson Cancer Center