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Sarah,

Both TIRF and parfocality will depend on correction for chromatic
aberrations (at least a tiny bit for the latter and maybe more for
TIRF). For TIRF, the "critical" angle is a function of the lens and
medium RI. If your objective is not perfectly corrected chromatically
over all wavelengths and/or the medium RI is sensitive to wavelength
(which it is), then the critical angle for different lines will be
different and require adjustment of the laser angle for each line to
get TIRF to function. As we all know, not all 1.45 or 1.6 NA
objectives perform alike.

As Michael points out, at 440 nm he gets good performance but must
change the angle when he excites at 488 nm and longer. For a
different objective, different angles and performance are to be
expected at different wavelengths.

Mario

>Search the CONFOCAL archive at
>http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>
>At 12:25 PM 02/09/05 -0500, you wrote:
>>Search the CONFOCAL archive at
>>http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>>Thanks for the info George.  You bring up a good point with the
>>parfocality issue.  I'm actually more interested in TIRF applications, so
>>it makes me wonder if parfocality will be more or less of an issue for TIRF?
>>Sarah
>
>
>The angle for TIRF varies with wavelength (at least with the Olympus N.A.
>1.45 objective).  We have a 440nm diode laser which works great for CFP,
>but when we want to use excitation at 488 nm we need to change the
>angle.  Often we can get away with one angle for 488 and 543, but even here
>tweaking at 543 improves the TIRF.
>
>
>
>____________________________________________________________________________
>Michael Cammer   Analytical Imaging Facility   Albert Einstein Coll. of Med.
>Jack & Pearl Resnick Campus      1300 Morris Park Ave.     Bronx, NY  10461
>(718) 430-2890       Fax:  430-8996      URL:  http://www.aecom.yu.edu/aif/
>   **This electronic transmission contains information that is privileged.**


--
________________________________________________________________________________
Mario M. Moronne, Ph.D.
NanoMed Technologies LLC
President and CTO
ph (510) 528-2400
FAX (510) 528-8076
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94706

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