[ome-users] Increase in intensity in every second frame in Leica lif files

[Melissa Linkert]

Hi Pascal,

> I just uploaded the zip folder "lif file problem" containing the
> four files on the specified website.
> 
> I am curious what the problem is.

Thank you for uploading an example dataset.

The problem is that the reported order of channels and timepoints does
not match the actual order.  We are reviewing a fix for the problem
here:

https://github.com/openmicroscopy/bioformats/pull/804

Once that shows as being closed, enabling the "Bio-Formats 5" update
site (in Fiji) or downloading the latest trunk build (in ImageJ) from:

http://downloads.openmicroscopy.org/bio-formats/5.0.0-beta1/

should solve the problem.

Regards,
-Melissa

On Thu, Nov 21, 2013 at 03:33:34PM +0100, Pascal Lorentz wrote:
> Dear Curtis
> 
> Thanks a lot for your reply.
> I just uploaded the zip folder "lif file problem" containing the
> four files on the specified website.
> 
> I am curious what the problem is.
> 
> Best regards
> 
> Pascal
> 
> Am 21.11.2013 10:18, schrieb Curtis Rueden:
> >Hi Pascal,
> >
> >> I also tried with Fiji. I updated Fiji through the menu and also
> >> manually updated the loci plugin there. I am not sure if it takes the
> >> trunk build in that case.
> >
> >Fiji ships with the 4.4.9 release of Bio-Formats. If you want to
> >enable 4.4.x or 5.x development builds, you can use the
> >"Bio-Formats 4" or "Bio-Formats 5" update sites. For more details,
> >please see the instructions at:
> >
> >http://fiji.sc/Bio-Formats#Daily_builds
> >
> >With Fiji, it should not be necessary to download loci_tools.jar
> >manually, nor to ever run the "Update LOCI Plugins" command (that
> >command is disabled in Fiji anyway).
> >
> >> Anyway it is always the same. We see the change in intensity in every
> >> second time frame.
> >
> >Indeed, it sounds like a bug.
> >
> >> I can provide you with 4 files but I need an ftp access
> >
> >Thanks. If you ZIP up the four files, you can upload them using this link:
> >
> >http://qa.openmicroscopy.org.uk/qa/upload/
> >
> >Unless your archive is >500MB? In which case, let us know and we
> >will provide alternative upload instructions off-list.
> >
> >Regards,
> >Curtis
> >
> >
> >On Thu, Nov 21, 2013 at 2:38 AM, Pascal Lorentz
> ><Lorentz.Pascal at gmail.com <mailto:Lorentz.Pascal at gmail.com>>
> >wrote:
> >
> >    Dear Bio Formats team
> >
> >    First of all I would like to thank you for your impressive work.
> >    This plugin is really great.
> >
> >    We came across a problem yesterday.
> >    We have a lif file that has been acquired with a Leica confocal
> >    SP5 with the LAS AF Software Version 2.7.3. It is a timelaps
> >    dataset with 12 timepoints in 2 channels. If we look at these
> >    cells in the Leica software the intensity over time stays in the
> >    same range.
> >    If we open this dataset as a hyperstack with the bio formats
> >    importer we see an increase in intensity in every second time
> >    frame. It is not just a display effect since we can really measure
> >    the intensity change.
> >    As a work around we exported single tifs from the lif file out of
> >    the Leica software. So we end up with 24 images for the 12
> >    timepoints for the two channels. If we then import these images as
> >    an image sequence and if we measure the exact same area, the
> >    intensity stays in a similar range over the whole timelaps.
> >
> >    I can provide you with 4 files but I need an ftp access to upload
> >    them:
> >    1. The original lif file “Series 7.lif”
> >    2. The image sequence after exporting from the Leica Software and
> >    importing as sequence in ImageJ saved as tiff “image_sequence.tif”
> >    3. The results table of the intensity measurement of an area of
> >    the lif dataset opened with the loci importer
> >    “Results_loci_import.xls.
> >    4. The results table of the intensity measurement of the same area
> >    as in point 3 after importing the single tifs as an image sequence.
> >
> >    My user saw that problem on a Mac with Fiji but we also saw it on
> >    my PC as well.
> >    I downloaded the latest stable release from
> >    http://downloads.openmicroscopy.org/bio-formats/4.4.9/#tools
> >    I also tried to update the plugin to the trunk build with Plugins
> >    -> Loci -> Update loci plugins but it always fails with the
> >    message “an error occurred while downloading the plugins”
> >    I did all that that with ImageJ 1.48g
> >    I also tried with Fiji. I updated Fiji through the menu and also
> >    manually updated the loci plugin there. I am not sure if it takes
> >    the trunk build in that case.
> >    Anyway it is always the same. We see the change in intensity in
> >    every second time frame.
> >    I also tried an older version of the loci plugin from 2010 but it
> >    shows the same effect.
> >
> >    We never so that problem with any lif files before. But our
> >    acquisition software has been updated a few weeks ago and I am not
> >    sure if Leica changed something on the file format.
> >
> >    So I was using ImageJ 1.48g with the loci plugin 4.4.9 on a
> >    windows 7 64bit machine with Java Version 7 Update 45 (build
> >    1.7.0_45-b18).
> >
> >    We would be more than happy if you can have a look at our problem.
> >    If you need more information just let me know.
> >
> >
> >    Many thanks and best regards
> >
> >    Pascal
> >
> >    _______________________________________________
> >    ome-users mailing list
> >    ome-users at lists.openmicroscopy.org.uk
> >    <mailto:ome-users at lists.openmicroscopy.org.uk>
> >    http://lists.openmicroscopy.org.uk/mailman/listinfo/ome-users
> >
> >
> 

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