*****
To join, leave or search the confocal microscopy listserv, go to:
http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
*****
Hi Niko
we had a similar problem but additionally observe a pattern of a certain
frequency bands which are more evident when we upgrade to increase the
sampling frequency.
after several trials we identified that the dichroic mirror used to reflect
the laser ablation was defective, it is easy to identify because if you can
see the lines and slightly move the mirror you can see how are you moving
lines.
hope will be of your help
best regards
Jorge
2013/8/22 Nic Cade <[log in to unmask]>
> *****
> To join, leave or search the confocal microscopy listserv, go to:
> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> *****
>
> Hi Niko,
>
> Which microscope system do you have this problem with? Do you also see a
> corresponding change in focus?
>
> We have a very similar problem with our 3i spinning disk system where we
> see a
> periodic oscillation in lateral position, ~250nm every 60s. We think we
> have
> narrowed it down to the heater for the incubation chamber (Okolab) which
> also
> switches on and off with a period of 60s. Another group here with the same
> system have found exactly the same thing; turning off the heater stopped
> the
> oscillations.
> I'm guessing that in your case the oscillations might be large enough to
> move
> your sample through the focal plane, but which wouldn't affect any
> featureless
> background. You could test this with some fluorescent beads if you haven't
> done so already.
>
> Cheers
> Nic
>
> On Fri, 16 Aug 2013 17:37:32 +0200, Niko Ehrenfeuchter
> <[log in to unmask]> wrote:
>
> >*****
> >To join, leave or search the confocal microscopy listserv, go to:
> >http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> >*****
> >
> >Dear all,
> >
> >we're recently facing a very strange behavior on one of our spinning
> >disk systems. One of our users noticed a repeated fluctuation in
> >recorded timelapse experiments (this was a FRAP experiment, but we don't
> >think that's related to FRAP).
> >
> >The timelapse was recorded every 6 seconds, and after background
> >subtraction a repeating drop of signal is visible in the intensity plot
> >(please find the corresponding graph under [1], see below). The
> >fluctuation is also visible by eye when scrolling through the timelapse.
> >
> >We noticed a frequency of 66 seconds and therefore think we can exclude
> >any external vibrations as source of this problem (plus the system is
> >mounted on a Melles Griot optical table). On some experiments the
> >oscillation almost follows a sine-like function, on others (like the
> >mentioned one) it looks more rectangular with a very rapid drop and
> >raise. Also, the frequency does *not* change if we double the
> >acquisition frequency, it's still on 66 seconds when acquiring every 3
> >seconds.
> >
> >One very weird thing is the stability of the background. The plots from
> >above show the intensity in selected ROI's over cells. The same plot for
> >an "empty" region in the sample shows a more or less constant signal (at
> >least we could figure a pattern in it, even after normalization and
> >using a big scaling factor to visualize the deltas).
> >
> >Has anyone of you come across a similar behavior?
> >
> >Any ideas about this would be highly appreciated!
> >
> >Many Thanks,
> >Niko
> >
> >[1]
> >
> https://www.biozentrum.unibas.ch/fileadmin/redaktion/Forschung/Research_Gr
> oups/IMCF/3i_intensity_fluctuation.png
> >
> >--
> >Niko Ehrenfeuchter
> >Imaging Core Facility
> >Kragenbau, Room G1055
> >Biozentrum, University of Basel
> >Klingelbergstrasse 50/70
> >CH-4056 Basel (Switzerland)
> >
> >Office: +41 (61) 26 72673
> >Email: [log in to unmask]
>
--
Jorge Toledo H.
Ph.D. student
Biomedical Neuroscience Institute (BNI)
Laboratory of Scientific Image Analysis (SCIAN-Lab)
Faculty of Medicine
University of Chile
www.scian.cl | www.couvelab.org
|
