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Hi Holly and list,
the scan head is controlled by an external computer, that is correct.
Using Nobody would use Windows for this task, also not for a point
scanner, if you see the time scale for the x/y scan mirrors. Time steps
for Windows and Mac OS are in the range of 60 ms, if I remember correct.
Unfortunately companies think that customers only buy systems that run
Windows. One won't get this problem with Linux (but maybe others). These
external "real-time computers" are usually controlled by Linux.
The faulty timing in the final image is added on the way from the external
computer to the Windows machine, since incoming data appears faster than
the OS can stamp it.
Now when it comes to triggering, as you plan it, Holly, I am not sure
which computer controls and handles this. In principle this needs to be
done directly via the RTC, but I recommend to get in touch with your local
Zeiss application specialist.
Michael
> Search the CONFOCAL archive at
> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>
> Hi, Craig & Geoff & Everyone -
>
>
>
> The 5-LIVE has a "real-time computer" or something it calls a real-time
> computer that the Windows box talks to. I would assume all the timing and
> handling is done with the real-time computer, but maybe it's not.
>
>
>
> The 5-LIVE is capable of 120 frames per second, but maybe not accurately?
> Even with a dedicated real-time computer? Or is there something not
> working
> correctly on our system?
>
>
>
> Thanks for this great discussion!
>
> -Holly
>
> __________________
>
> Holly L. Aaron
>
> CRL Molecular Imaging Center
>
> http://imaging.berkeley.edu
>
> _____
>
> From: Confocal Microscopy List [mailto:[log in to unmask]] On
> Behalf Of Craig Brideau
> Sent: Friday, July 11, 2008 1:56 PM
> To: [log in to unmask]
> Subject: Re: Zeiss 5-LIVE and Timing Issues
>
>
>
> Search the CONFOCAL archive at
> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal Try doing the same
> thing with a different microscope but similar image parameters; same bit
> depth per pixel, number of pixels, and acquisition times. I'll bet you
> see
> similar performance issues. We have some homebrew microscopes that I have
> worked with where we encountered operating system timing problems. I
> actually had to offload a lot of timing stuff, like the pixel clock
> generation, to external hardware (pre-programmed timer/counter modules
> with
> their own hardware clocks) because windows cannot generate a reliable
> timing
> signal in the ms range due to the operating system's event handling
> limitations. Again, you either need dedicated external hardware or a
> real-time OS to get reliable control over timing for small timing
> intervals,
> and Windows (and actually most desktop OSes) simply don't provide this.
>
> Craig
>
>
>
> On Fri, Jul 11, 2008 at 2:50 PM, Lambright, Geoffrey
> <[log in to unmask]> wrote:
>
> Search the CONFOCAL archive at
> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>
> Hi Craig, and list,
>
>
>
> So if it is the case of the computer hardware "shuffling" the data that is
> causing the imprecision in the data timing, does that mean that the timing
> of the actual image acquisition is precise and the variance that Holly
> sees
> comes from the computer's ability to process and record the captured data?
> Any idea how one could check for that if that was the case?
>
>
>
> Geoff
>
>
>
> From: Confocal Microscopy List [mailto:[log in to unmask]] On
> Behalf Of Craig Brideau
> Sent: Friday, July 11, 2008 1:03 PM
> To: [log in to unmask]
> Subject: Re: Zeiss 5-LIVE and Timing Issues
>
>
>
> Search the CONFOCAL archive at
> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal Hi Holly! This
> may
> be dependent on the computer hardware running the `scope. If it's
> Windows,
> then all bets are off for timing when you get down to the low hundreds or
> tens of ms. Basically the OS can't reliably shuffle data around fast
> enough
> at those speeds; you need a real-time operating system for that. In the
> case of most multitasking operating systems no single process can 'bogart'
> the CPU, and the time to switch between process threads can vary. What
> this
> boils down to is that the computer will take its own sweet time handling
> the
> data coming in from your microscope depending on what else is going on
> with
> the computer and the time scale we are talking about.
>
> Craig
>
>
>
> On Fri, Jul 11, 2008 at 12:14 PM, Holly Aaron <[log in to unmask]> wrote:
>
> Search the CONFOCAL archive at
> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>
> Dear Confocal Community -
>
>
>
> This question may be very specific to the Zeiss community and even more
> specific to the 5-LIVE users out there.
>
> We find the 5-LIVE to be unreliable/unpredictable/unrepeatable in time
> intervals shorter than 100msec. By this I mean that if we set up a
> time-series in which an image should be taken every 500ms (let's say the
> time required for the image is 30ms), it works well: an image is in fact
> taken every 500ms. However, if we then decide we want a shorter interval,
> 100ms or less (which should be fine given only 30ms for each frame), the
> images are taken at random times, some greater than 100ms, some less:
>
>
>
>
> Image #
>
> Time Expected
>
> Time Actual
>
>
> 1
>
> 0
>
> 0
>
>
> 2
>
> 0.1
>
> 0.0987
>
>
> 3
>
> 0.2
>
> 0.1998
>
>
> 4
>
> 0.3
>
> 0.3009
>
>
> 5
>
> 0.4
>
> 0.4008
>
>
>
> This gets worse when we go to shorter and shorter intervals, for example,
> 50ms:
>
>
> Image #
>
> Time Expected
>
> Time [ s]
>
>
> 1
>
> 0
>
> 0
>
>
> 2
>
> 0.05
>
> 0.0209
>
>
> 3
>
> 0.1
>
> 0.1064
>
>
> 4
>
> 0.15
>
> 0.1273
>
>
> 5
>
> 0.2
>
> 0.2104
>
>
>
> And even worse for shorter.
>
>
>
> I am wondering: Does anyone else see this phenomenon? Have you been able
> to
> get around it? We are doing electrophysiology in concert with imaging and
> timing is crucial. So far we have not been able to set it up to trigger
> each
> image because that takes too long and is even less predictable. We would
> be
> very grateful for anyone who has found a work-around for this problem.
>
>
>
> Thank you!
>
> __________________
>
> Holly L. Aaron
>
> Molecular Imaging Center
>
> Cancer Research Laboratory
>
> University of California Berkeley
>
> 447 LSA #2751
>
> Berkeley, CA 94720-2751
>
> 510.642.2901
>
> 510.642.5741 fax
>
> [log in to unmask]
>
> http://imaging.berkeley.edu
|
