MetaMorph Stack: issue with OME metadata (bioformats and OMERO)

Dear all,

A user at our facility has recorded time-laps sequences in *.nd and *.stk format with our Visitron Spinning Disk microscope (.nd and associated .stk files), and reported that some metadata was not correct while opened with Bio-Formats or imported in OMERO. The files are in the MetaMorph Stack STK file format.

  • The folder 60x_2well_all100_G50ms_C100ms contain data that was acquired by setting the exposure time for the 488nm laser to 50ms and the exposure time for the 640nm laser to 100ms. However, from the OME metadata, it appears that all channels have the same exposure time (50ms).
  • Also, it incorrectly assigns the emission wavelength to the laser wavelength (the same 525nm for the two lasers, sdcGFP and sdcCy5.
  • Also, in the OME metadata, DeltaT is set to 0 starting from the 6th timepoint, although the user have acquired images every 20s.

from the OME metadata (Bioformats):


from OMERO.web:

Could someone help me and my colleague @oburri with that, and tell us more about the Bioformats STK reader?

All the best,
Claire

Hi @stoffelc, there have a few MetaMorph fixes in recent Bio-Formats releases specifically around the parsing of laser metadata so it may be worth checking to see how those changes impact the files.

Do you know which Bio-Formats version you were using when testing? Updating the version in FIJI to the latest 6.6.0

Alternatively if you download the latest command line tools from Bio-Formats Downloads | Open Microscopy Environment (OME) and then run the below command, how does that compare with what you are currently seeing?

showinf -nopix -omexml path/to/myStack.nd

1 Like

Hi @dgault,

Thank you very much for your answer. I have the latest version of FIJI, and running the showinf -nopix -omexml path/to/myStack.nd command gave me the same metadata:


   <Plane DeltaT="0.0" DeltaTUnit="s" ExposureTime="0.05" ExposureTimeUnit="s" PositionX="-22699.8" PositionXUnit="reference frame" PositionY="-3262.0" PositionYUnit="reference frame" PositionZ="166.25" PositionZUnit="reference frame" TheC="0" TheT="0" TheZ="0"/>
         <Plane DeltaT="0.075" DeltaTUnit="s" ExposureTime="0.05" ExposureTimeUnit="s" PositionX="-22699.8" PositionXUnit="reference frame" PositionY="-3262.0" PositionYUnit="reference frame" PositionZ="168.125" PositionZUnit="reference frame" TheC="0" TheT="0" TheZ="1"/>
         <Plane DeltaT="0.15" DeltaTUnit="s" ExposureTime="0.05" ExposureTimeUnit="s" PositionX="-22699.8" PositionXUnit="reference frame" PositionY="-3262.0" PositionYUnit="reference frame" PositionZ="170.0" PositionZUnit="reference frame" TheC="0" TheT="0" TheZ="2"/>
         <Plane DeltaT="0.226" DeltaTUnit="s" ExposureTime="0.05" ExposureTimeUnit="s" PositionX="-22699.8" PositionXUnit="reference frame" PositionY="-3262.0" PositionYUnit="reference frame" PositionZ="171.875" PositionZUnit="reference frame" TheC="0" TheT="0" TheZ="3"/>
         <Plane DeltaT="0.3" DeltaTUnit="s" ExposureTime="0.05" ExposureTimeUnit="s" PositionX="-22699.8" PositionXUnit="reference frame" PositionY="-3262.0" PositionYUnit="reference frame" PositionZ="173.75" PositionZUnit="reference frame" TheC="0" TheT="0" TheZ="4"/>
         <Plane DeltaT="0.0" DeltaTUnit="s" ExposureTime="0.05" ExposureTimeUnit="s" TheC="1" TheT="0" TheZ="0"/>
         <Plane DeltaT="0.0" DeltaTUnit="s" ExposureTime="0.05" ExposureTimeUnit="s" TheC="1" TheT="0" TheZ="1"/>
         <Plane DeltaT="0.0" DeltaTUnit="s" ExposureTime="0.05" ExposureTimeUnit="s" TheC="1" TheT="0" TheZ="2"/>
         <Plane DeltaT="0.0" DeltaTUnit="s" ExposureTime="0.05" ExposureTimeUnit="s" TheC="1" TheT="0" TheZ="3"/>
         <Plane DeltaT="0.0" DeltaTUnit="s" ExposureTime="0.05" ExposureTimeUnit="s" TheC="1" TheT="0" TheZ="4"/>
         <Plane DeltaT="0.0" DeltaTUnit="s" ExposureTime="0.05" ExposureTimeUnit="s" TheC="0" TheT="1" TheZ="0"/>
         <Plane DeltaT="0.0" DeltaTUnit="s" ExposureTime="0.05" ExposureTimeUnit="s" TheC="0" TheT="1" TheZ="1"/>
         <Plane DeltaT="0.0" DeltaTUnit="s" ExposureTime="0.05" ExposureTimeUnit="s" TheC="0" TheT="1" TheZ="2"/>
         <Plane DeltaT="0.0" DeltaTUnit="s" ExposureTime="0.05" ExposureTimeUnit="s" TheC="0" TheT="1" TheZ="3"/>
         <Plane DeltaT="0.0" DeltaTUnit="s" ExposureTime="0.05" ExposureTimeUnit="s" TheC="0" TheT="1" TheZ="4"/>
         <Plane DeltaT="0.0" DeltaTUnit="s" ExposureTime="0.05" ExposureTimeUnit="s" TheC="1" TheT="1" TheZ="0"/>
         <Plane DeltaT="0.0" DeltaTUnit="s" ExposureTime="0.05" ExposureTimeUnit="s" TheC="1" TheT="1" TheZ="1"/>
         <Plane DeltaT="0.0" DeltaTUnit="s" ExposureTime="0.05" ExposureTimeUnit="s" TheC="1" TheT="1" TheZ="2"/>
         <Plane DeltaT="0.0" DeltaTUnit="s" ExposureTime="0.05" ExposureTimeUnit="s" TheC="1" TheT="1" TheZ="3"/>
         <Plane DeltaT="0.0" DeltaTUnit="s" ExposureTime="0.05" ExposureTimeUnit="s" TheC="1" TheT="1" TheZ="4"/>
         <Plane DeltaT="0.0" DeltaTUnit="s" ExposureTime="0.05" ExposureTimeUnit="s" TheC="0" TheT="2" TheZ="0"/>
         <Plane DeltaT="0.0" DeltaTUnit="s" ExposureTime="0.05" ExposureTimeUnit="s" TheC="0" TheT="2" TheZ="1"/>
         <Plane DeltaT="0.0" DeltaTUnit="s" ExposureTime="0.05" ExposureTimeUnit="s" TheC="0" TheT="2" TheZ="2"/>

and

  <Channel EmissionWavelength="525.0" EmissionWavelengthUnit="nm" ID="Channel:2:0" Name="sdcGFP" SamplesPerPixel="1">
            <LightSourceSettings ID="LightSource:0:4" Wavelength="525.0" WavelengthUnit="nm"/>
            <DetectorSettings Binning="1x1" Gain="0.0" ID="Detector:0:0"/>
            <LightPath/>
         </Channel>
         <Channel EmissionWavelength="525.0" EmissionWavelengthUnit="nm" ID="Channel:2:1" Name="sdcCy5" SamplesPerPixel="1">
            <LightSourceSettings ID="LightSource:0:5" Wavelength="525.0" WavelengthUnit="nm"/>
            <DetectorSettings Binning="1x1" Gain="0.0" ID="Detector:0:0"/>
            <LightPath/>
         </Channel>

Best,
Claire

Hi Clarie,

just to be on the safe side, can you confirm which version of Bio-Formats is in your FIJI via Help -> About Plugins -> Bio-Formats Plugins?

Thanks!
~Josh

Hi @joshmoore,

Version is 6.6.1:

best,
Claire

Thanks Claire, would it be possible to share the dataset (nd file and any associated stk or tiff) to a public location so I can further investigate the issue? If you need an upload location we would recommend using Zenodo.

Hi David,
Sorry it may not have been obvious, but I shared the dataset with a switchdrive link here:

all the best,
Claire

Thanks Claire, sorry I missed the link earlier. Those issues are Bio-Formats bugs and I have opened a Bio-Formats GitHub Issue to track the bugs: MetaMorph: Incorrect parsing of laser metadata · Issue #3662 · ome/bioformats · GitHub

1 Like

No worries, thank you for having opened a GitHub Issue to track those Bio-Formats bugs!