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

[Pascal Lorentz]

Dear Melissa

Thank you very much. I really appreciate your help.

Best regards

Pascal

Am 22.11.2013 03:27, schrieb 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
>>>
>>>
>> _______________________________________________
>> ome-users mailing list
>> ome-users at lists.openmicroscopy.org.uk
>> http://lists.openmicroscopy.org.uk/mailman/listinfo/ome-users