Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

Make sure that you are saving and updating the config after every change
and before you open the MULTITIME macro. Also, on our system you have to
make sure that the stage is at the "correct" focal plane when starting a
z-stack. the system doesn't seem to know where to start, so it just
starts wherever you happen to be focused. I know you aren't doing a
z-stack, but if you are moving the focus plane after saving the config
it could cause problems.

I dread people asking me to use Multitime because things like this seem
to happen all the time.

Good luck,
Robert


Yi Zheng wrote:
> Search the CONFOCAL archive at
> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>
> Dear all,
>
> I am a newbie to Zeiss LSM510.
>
> Recently I tried to run a multiple locations time series macro on LSM510.
> The protocol I followed is the "Zeiss LSM 510 META NLO User's Manual" from 
> Fred Hutchinson Cancer Research Center which I got online.
>
> After mark 4 different location and load my configuration, I did these step as 
> follows:
>
> 6. Start Multi-Time Macro (Main Menu>MACROS>MultiTime"R")
> 7. Click "Refresh" button.
> 8. Load Multi-Track configuration from pull-down menu in Multi-Time
> window (Config, Time-Interval and Bleach), (e.g. aa1)
> 9. Make sure "number of scans" is set to "0", and z-stack is checked in the
> "Config, Time-Interval and Bleach" panel.(I did not check the z-stack because 
> I only have one z-stack)
> 10. Under "List of Blocks", set number of repetitions to "0"(I changed this 
> number to 3 later in step 15.1)
> 11. Make sure autofocus is NOT checked, and bleach is NOT checked.
> 12. Click "Multiple Location" button on top of Multi-Time window.
> 13. Verify that the list of locations in the Multi-Time series window menu
> matches the list in the stage control panel. If not, click Refresh.
> 14. Click "Options" (in Multi_Time window); Check "Keep Final Image
> Open" and "Save Final Image"
> 15.1. Under List of Blocks, enter the number of Block repetitions (e.g. 3, for
> three time-points)
> 15.2. Under Block 1 parameters, enter the wait interval.
> 16. Click "Start Time".
>
> But after all these, no images were collected except the black one and 
> warning message as "Pinhole PH2 is not adjusted".  I also notice that in 
> configuration control panel, under list of tracks, the channels info changed to 
> another laser momentarily (it switched from this to my previous setting 
> continuously). In the end, I had to stop the recording manualy and no image 
> was saved.
>
> I have attached three pics of my setting screen, and would you guys please 
> check my protocol and let me know which step I made a mistake?
>
> Here is the link
> [IMG]http://i143.photobucket.com/albums/r138/qqbmwcichlid/macro-1.jpg
> [/IMG]
>
> [IMG]http://i143.photobucket.com/albums/r138/qqbmwcichlid/macro-3.jpg
> [/IMG]
>
> [IMG]http://i143.photobucket.com/albums/r138/qqbmwcichlid/macro-2.jpg
> [/IMG]
>
> Thanks in advance for any suggestions.
>
>
> Yi
>   



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