http://confocal-microscopy-list.275.s1.nabble.com/Re-Objective-lens-chromatic-aberration-tp5929621p5929995.html
lens was at fault. I must admit I don't see how with rear aperture
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> Hi Michael,
>
> There are a few factors that need to be compromised in TIRF. 1)
> The TIRF
> penetration depth is dependent upon the wavelength of the incident
> illumination, the angle of incidence, and the refractive indices of
> the
> media at the interface. 2) An apochromatic lens brings lasers of three
> wavelengths on to the same focal plane, but may not to the same
> incident
> angle. Therefore, the incident angles for the three lasers should
> be able
> to be adjusted independently to get optimal TIRF images.
>
> You may want to take your sample off first, to see how different the
> angles
> of the three lasers projecting to the wall, to get an idea of the
> incident
> angles of your lasers coming out of the objective.
>
> Regards
>
> Gary G Li, PhD
>
>
> On Sun, Jan 16, 2011 at 1:02 PM, Cammer, Michael <
[hidden email]
>> wrote:
>
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>> I originally posted the message below to the microscopy listserv but
>> reviewing the discussion here on chromatic aberration, I thought it
>> would be
>> apropos here too even though it is not really regarding confocal.
>>
>> A few years ago we were having problems with the first commercial
>> Olympus
>> TIRF system because we could not get consistent evanescent waves
>> with the
>> one angle adjustment with the laser lines we had from 405 to 568 nm
>> that
>> were delivered via a single fiber (it was worse when we later added
>> a 633 nm
>> laser). I suggested we pump each laser in through a separate path
>> that
>> could be angled independently. We didn't build it, but I think
>> Olympus now
>> sells a TIRF system that does this.
>>
>> Another issue is that when I first heard about TIRF maybe 15 years
>> ago, it
>> was introduced as a ring illumination at the outer edge of the back
>> aperture, not as a single point or crescent at the periphery on
>> only one
>> side. A ring, or at least a series of points around the periphery,
>> seems
>> like a better way to provide a uniform field due to aberrations from
>> coherent light in the imperfect optics. Any thought on this?
>>
>> Sincerely,
>>
>> Michael
>>
>> -----------------------ORIGINAL
>> MESSAGE-------------------------------
>> We have the Nikon TIRF system and have three laser lines
>> going into the TIRF arm via a single fiber. When we project through
>> the 100X objective through the sample onto the wall we see that the
>> lines go through the sample at different angles. (You can see a
>> picture of the projection at approx 45 degrees at
>>
http://www.flickr.com/photos/mcammer/5359189090/ .) It is also
>> noticeable in the TIRF images that the field depth is different for
>> each wavelength. Is this unavoidable due to the different
>> wavelengths or is it possible to align the optics better so these
>> spots would be more coincident?
>>
>>
>>
>> _________________________________________
>> Michael Cammer, Assistant Research Scientist
>> Skirball Institute of Biomolecular Medicine
>> Lab: (212) 263-3208 Cell: (914) 309-3270
>>
>>
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