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Andreas, can expound on the sentence below? I'm not sure what you mean
exactly.
Thank you!
"In strongly scattering samples like lymph nodes the signal decreases so
rapidly with depth that you hardly notice a difference in penetration
depth when adjusting the pre-chirp".
Brian D Armstrong PhD
Light Microscopy Core Manager
Beckman Research Institute
City of Hope
Dept of Neuroscience
1450 E Duarte Rd
Duarte, CA 91010
626-256-4673 x62872
http://www.cityofhope.org/research/support/Light-Microscopy-Digital-Imag
ing/Pages/default.aspx
-----Original Message-----
From: Confocal Microscopy List [mailto:[log in to unmask]]
On Behalf Of Andreas Bruckbauer
Sent: Thursday, November 18, 2010 11:09 PM
To: [log in to unmask]
Subject: Re: MaiTai DeepSee adjustments on multiphoton microscope
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Hi Michael,
i tried a quantification at 800 nm excitation 450 - 500 micrometer into
fixed lymph nodes using the 25x Olympus objective. Over the range of the
deepsee compensation i found a change of a factor two in fluorescence.
One thing is that i could not set the compensation to zero with our
deepsee model so i don't know what the fluorescence would be without
deepsee. Anyhow with a good estimate of the GDD of the system from the
deepsee motor position one can calculate how much this would broaden the
100 fs pulse out of the MaiTai and for our system this is about 220 fs.
so you would have to use 1.5x more laser power to get the same signal
without pre-chirp.
In my experience you don't need to optimise for different samples as the
less than one mm depth into the sample is insignificant compared to the
thickness of the optics and GVD of an AOM. There is a slight difference
for different objectives though (more or less glass). My conclusion was
that for live samples the pre-chirp is definetly a benefit because you
can use lower laser power. In strongly scattering samples like lymph
nodes the signal decreases so rapidly with depth that you hardly notice
a difference in penetration depth when adjusting the pre-chirp.
best wishes
Andreas
-----Original Message-----
From: Cameron Nowell <[log in to unmask]>
To: [log in to unmask]
Sent: Thu, 18 Nov 2010 21:58
Subject: Re: MaiTai DeepSee adjustments on multiphoton microscope
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Hi Michael,
We have the DeepSee on our MaiTai and have found that it doesn't do a
lot on our system. At extreme depths, say greater than 500um you can get
a small enhancement in signal strength and quality but it is only very
minor. It may give you a few extra microns penetration but thats about
it.
It really depends on how much the optics of your system are
dispersing/chirping the beam. If they are not altering it much then
pre-chirping it will not do a lot.
This is all based on qualitative assessment, i have not quantitated any
of this. Also we have recently received a new Olympus 25x NA1.05
objective specifically designed for MP imaging. I have not seen how this
performs yet with the DeepSee.
Cheers
Cam
Cameron J. Nowell
Microscopy Manager
Centre for Advanced Microscopy
Ludwig Institute for Cancer Research
PO Box 2008
Royal Melbourne Hospital
Victoria, 3050
AUSTRALIA
Office: +61 3 9341 3155
Mobile: +61422882700
Fax: +61 3 9341 3104
Facility Website
-----Original Message-----
From: Confocal Microscopy List [mailto:[log in to unmask]]
On Behalf Of Cammer, Michael
Sent: Friday, 19 November 2010 4:08 AM
To: [log in to unmask]
Subject: MaiTai DeepSee adjustments on multiphoton microscope
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Dear microscopists,
I'm planning to go through a regimen of adjusting the MaiTai DeepSee
on a variety of samples with different optics to see what changes there
are in depth penetration (and, maybe, preservation of biological
activity).
Does anybody have experience with this? If so, have you found that the
DeepSee provides a noticeable practical benefit over the MaiTai without
it?
Any comments welcome and please feel free to contact me offline at
[log in to unmask]
Thank you!
Sincerely,
Michael C.
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