Organizing a live-cell imaging core

[Martin Wessendorf-2]

Dear List--

For those of you who have successful multi-photon live-cell imaging
cores: how have you set up your facilities?  If different users have
different requirements (e.g. upright vs. inverted stand; temperature/CO2
control; ability to record electrophysiologically; video-rate scanning
vs. slower scanning), how do you accommodate them all?  Are there some
instruments or configurations that are significantly more flexible
and/or that provide more capabilities for the money spent?  Are there
some types of experiments that so problematic that they're not worth
attempting to support?

Thanks.  Responses on- or off-list are welcome.

Martin Wessendorf
--
Martin Wessendorf, PhD   [phone] (office)
Associate Professor      [phone] (lab)
Dept Neuroscience      [phone] (FAX)
Univ Minnesota        e-mail: martinw(at)umn.edu

[Csúcs Gábor]

Dear Martin,

I will shortly describe our systems, and afterwards explain you why I
wouldn't do it once more... :-)))
We have on the same optical table a Zeiss Axisoscope FS (fixed stage)
and an Axiovert 200M sisntrument. The confocal system is an
LSM510Meta-NLO with a Coherent Chameleon laser mounted on the same
optical table. Our idea was to use the same confocal head on both stages
(with the Zeiss systems this is relatively easy). The light path of the
Chameleon was built in a way that can be directed either to the inverted
stage or to the upright one. So the idea was to have one confocal system
but change the stages depending on the application. Now, after a while
it turned out that this is unrealistic in a core facility. Although
mounting the confocal head is relatively easy, but re-aligning the
multi-photon laser requires much more care/time - so you'll never do it
on a daily base (even a weekly one is too much). So my current
suggestion for such a system would be to purchase one upright system and
one inverted one with a dedicated confocal head and sharing only the
multi-photon laser between them. Electrophysiology is a rather special
area with a somewhat tricky instrumentation (amplifiers etc.) - so I
would not really suggest to host it in a general microscopy core
facility. For enhanced flexibility on this system we use only a stage
incubator (which is easy to remove etc.) and not a typical box-type one.

Cheers   Gabor

> Dear List--
>
> For those of you who have successful multi-photon live-cell imaging
> cores: how have you set up your facilities?  If different users have
> different requirements (e.g. upright vs. inverted stand;
> temperature/CO2 control; ability to record electrophysiologically;
> video-rate scanning vs. slower scanning), how do you accommodate them
> all?  Are there some instruments or configurations that are
> significantly more flexible and/or that provide more capabilities for
> the money spent?  Are there some types of experiments that so
> problematic that they're not worth attempting to support?
>
> Thanks.  Responses on- or off-list are welcome.
>
> Martin Wessendorf

[Craig Brideau]

We had three microscopes attached to a single Tsunami ultrafast laser.
 We used polarizing beamsplitter cubes to route the power between the
three microscopes so that alignment was not an issue.  Rotating
waveplates sent the power from the laser to one or the other
microscope without having to change the alignment of any beam path.
One microscope was an upright (Nikon C1), the other was a modified
Nikon inverted (TE2000 I think) and the last was built from scratch.
The problem we encountered is that every user wanted to use a
different wavelength!  One researcher used 840nm all the time, while
another used 925nm.  As a result, they could not run their experiments
simultaneously.  In our new lab setup, we have one dedicated laser for
each scope to eliminate this problem.  If you must share a laser, use
the cube method for power-routing, and try to get your researchers to
use dyes that can be stimulated by the same wavelength!

Craig Brideau
Hotchkiss Brain Institute
University of Calgary


On Sun, Oct 25, 2009 at 3:29 PM, Gabor Csucs <[hidden email]> wrote:

> Dear Martin,
>
> I will shortly describe our systems, and afterwards explain you why I
> wouldn't do it once more... :-)))
> We have on the same optical table a Zeiss Axisoscope FS (fixed stage) and an
> Axiovert 200M sisntrument. The confocal system is an LSM510Meta-NLO with a
> Coherent Chameleon laser mounted on the same optical table. Our idea was to
> use the same confocal head on both stages (with the Zeiss systems this is
> relatively easy). The light path of the Chameleon was built in a way that
> can be directed either to the inverted stage or to the upright one. So the
> idea was to have one confocal system but change the stages depending on the
> application. Now, after a while it turned out that this is unrealistic in a
> core facility. Although mounting the confocal head is relatively easy, but
> re-aligning the multi-photon laser requires much more care/time - so you'll
> never do it on a daily base (even a weekly one is too much). So my current
> suggestion for such a system would be to purchase one upright system and one
> inverted one with a dedicated confocal head and sharing only the
> multi-photon laser between them. Electrophysiology is a rather special area
> with a somewhat tricky instrumentation (amplifiers etc.) - so I would not
> really suggest to host it in a general microscopy core facility. For
> enhanced flexibility on this system we use only a stage incubator (which is
> easy to remove etc.) and not a typical box-type one.
>
> Cheers   Gabor
>>
>> Dear List--
>>
>> For those of you who have successful multi-photon live-cell imaging cores:
>> how have you set up your facilities?  If different users have different
>> requirements (e.g. upright vs. inverted stand; temperature/CO2 control;
>> ability to record electrophysiologically; video-rate scanning vs. slower
>> scanning), how do you accommodate them all?  Are there some instruments or
>> configurations that are significantly more flexible and/or that provide more
>> capabilities for the money spent?  Are there some types of experiments that
>> so problematic that they're not worth attempting to support?
>>
>> Thanks.  Responses on- or off-list are welcome.
>>
>> Martin Wessendorf
>