Open Microscopy Environment

Hi,

I am new to the Bio-formats tool and I am trying to use it to import .vsi files in Fiji and/or MATLAB. However, whenever I try to import the image in either Fiji or MATLAB, I get a series of error messages (copied below). Any advice on how this can be solved?

Thanks in advance,

Stefanie

java.lang.NullPointerException
at java.lang.System.arraycopy(Native Method)
at loci.formats.in.CellSensReader.openBytes(CellSensReader.java:555)
at loci.formats.ChannelFiller.openBytes(ChannelFiller.java:156)
at loci.formats.ChannelFiller.openBytes(ChannelFiller.java:148)
at loci.formats.ChannelSeparator.openBytes(ChannelSeparator.java:198)
at loci.formats.ReaderWrapper.openBytes(ReaderWrapper.java:348)
at loci.formats.DimensionSwapper.openBytes(DimensionSwapper.java:249)
at loci.formats.ReaderWrapper.openBytes(ReaderWrapper.java:348)
at loci.formats.MinMaxCalculator.openBytes(MinMaxCalculator.java:269)
at loci.formats.MinMaxCalculator.openBytes(MinMaxCalculator.java:260)
at loci.formats.ReaderWrapper.openBytes(ReaderWrapper.java:334)
at loci.formats.ReaderWrapper.openBytes(ReaderWrapper.java:334)
at loci.plugins.util.ImageProcessorReader.openProcessors(ImageProcessorReader.java:186)
at loci.plugins.in.ImagePlusReader.readProcessors(ImagePlusReader.java:422)
at loci.plugins.in.ImagePlusReader.readPlanes(ImagePlusReader.java:387)
at loci.plugins.in.ImagePlusReader.readImage(ImagePlusReader.java:282)
at loci.plugins.in.ImagePlusReader.readImages(ImagePlusReader.java:243)
at loci.plugins.in.ImagePlusReader.readImages(ImagePlusReader.java:221)
at loci.plugins.in.ImagePlusReader.openImagePlus(ImagePlusReader.java:116)
at loci.plugins.in.Importer.readPixels(Importer.java:149)
at loci.plugins.in.Importer.run(Importer.java:86)
at loci.plugins.LociImporter.run(LociImporter.java:78)
at ij.IJ.runUserPlugIn(IJ.java:217)
at ij.IJ.runPlugIn(IJ.java:181)
at ij.Executer.runCommand(Executer.java:137)
at ij.Executer.run(Executer.java:66)
at java.lang.Thread.run(Thread.java:745)

Hi Stefanie,

Sorry to hear you are having problems. If possible would you be able to upload a sample file for testing to http://qa.openmicroscopy.org.uk/qa/upload/

Also which version of Bio-Formats have you been using? (The version can be found in FIJI under Help > About Plugins > Bio-Formats Plugins)

David Gault

Hi David,

Thanks for your response. Unfortunately I cannot share an image, because there is still some patient data visible on the images although we covered the patient information with labels.

I am using release 5.7.1 of the Bio-Formats Plugin.

I am copying the metadata of one of the images that I am trying to import below. I hope this can give some guidance on how to solve this.

Thanks for your help,

Stefanie

20x BitsPerPixel 8
20x DimensionOrder XYCZT
20x IsInterleaved true
20x IsRGB false
20x LittleEndian false
20x PixelType uint8
20x SizeC 3
20x SizeT 1
20x SizeX 35800
20x SizeY 20759
20x SizeZ 1
Image_T1.vsi #10 BitsPerPixel 8
Image_T1.vsi #10 DimensionOrder XYCZT
Image_T1.vsi #10 IsInterleaved true
Image_T1.vsi #10 IsRGB false
Image_T1.vsi #10 LittleEndian false
Image_T1.vsi #10 PixelType uint8
Image_T1.vsi #10 SizeC 3
Image_T1.vsi #10 SizeT 1
Image_T1.vsi #10 SizeX 948
Image_T1.vsi #10 SizeY 1896
Image_T1.vsi #10 SizeZ 1
Image_T1.vsi #11 BitsPerPixel 8
Image_T1.vsi #11 DimensionOrder XYCZT
Image_T1.vsi #11 IsInterleaved true
Image_T1.vsi #11 IsRGB false
Image_T1.vsi #11 LittleEndian false
Image_T1.vsi #11 PixelType uint8
Image_T1.vsi #11 SizeC 3
Image_T1.vsi #11 SizeT 1
Image_T1.vsi #11 SizeX 474
Image_T1.vsi #11 SizeY 948
Image_T1.vsi #11 SizeZ 1
Image_T1.vsi #12 BitsPerPixel 8
Image_T1.vsi #12 DimensionOrder XYCZT
Image_T1.vsi #12 IsInterleaved true
Image_T1.vsi #12 IsRGB false
Image_T1.vsi #12 LittleEndian false
Image_T1.vsi #12 PixelType uint8
Image_T1.vsi #12 SizeC 3
Image_T1.vsi #12 SizeT 1
Image_T1.vsi #12 SizeX 237
Image_T1.vsi #12 SizeY 474
Image_T1.vsi #12 SizeZ 1
Image_T1.vsi #14 BitsPerPixel 8
Image_T1.vsi #14 DimensionOrder XYCZT
Image_T1.vsi #14 IsInterleaved true
Image_T1.vsi #14 IsRGB false
Image_T1.vsi #14 LittleEndian false
Image_T1.vsi #14 PixelType uint8
Image_T1.vsi #14 SizeC 3
Image_T1.vsi #14 SizeT 1
Image_T1.vsi #14 SizeX 17900
Image_T1.vsi #14 SizeY 10380
Image_T1.vsi #14 SizeZ 1
Image_T1.vsi #15 BitsPerPixel 8
Image_T1.vsi #15 DimensionOrder XYCZT
Image_T1.vsi #15 IsInterleaved true
Image_T1.vsi #15 IsRGB false
Image_T1.vsi #15 LittleEndian false
Image_T1.vsi #15 PixelType uint8
Image_T1.vsi #15 SizeC 3
Image_T1.vsi #15 SizeT 1
Image_T1.vsi #15 SizeX 8950
Image_T1.vsi #15 SizeY 5190
Image_T1.vsi #15 SizeZ 1
Image_T1.vsi #16 BitsPerPixel 8
Image_T1.vsi #16 DimensionOrder XYCZT
Image_T1.vsi #16 IsInterleaved true
Image_T1.vsi #16 IsRGB false
Image_T1.vsi #16 LittleEndian false
Image_T1.vsi #16 PixelType uint8
Image_T1.vsi #16 SizeC 3
Image_T1.vsi #16 SizeT 1
Image_T1.vsi #16 SizeX 4475
Image_T1.vsi #16 SizeY 2595
Image_T1.vsi #16 SizeZ 1
Image_T1.vsi #17 BitsPerPixel 8
Image_T1.vsi #17 DimensionOrder XYCZT
Image_T1.vsi #17 IsInterleaved true
Image_T1.vsi #17 IsRGB false
Image_T1.vsi #17 LittleEndian false
Image_T1.vsi #17 PixelType uint8
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Image_T1.vsi #17 SizeT 1
Image_T1.vsi #17 SizeX 2238
Image_T1.vsi #17 SizeY 1298
Image_T1.vsi #17 SizeZ 1
Image_T1.vsi #18 BitsPerPixel 8
Image_T1.vsi #18 DimensionOrder XYCZT
Image_T1.vsi #18 IsInterleaved true
Image_T1.vsi #18 IsRGB false
Image_T1.vsi #18 LittleEndian false
Image_T1.vsi #18 PixelType uint8
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Image_T1.vsi #18 SizeT 1
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Image_T1.vsi #18 SizeY 649
Image_T1.vsi #18 SizeZ 1
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Image_T1.vsi #19 DimensionOrder XYCZT
Image_T1.vsi #19 IsInterleaved true
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Image_T1.vsi #19 LittleEndian false
Image_T1.vsi #19 PixelType uint8
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Image_T1.vsi #19 SizeT 1
Image_T1.vsi #19 SizeX 560
Image_T1.vsi #19 SizeY 325
Image_T1.vsi #19 SizeZ 1
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Image_T1.vsi #2 DimensionOrder XYCZT
Image_T1.vsi #2 IsInterleaved true
Image_T1.vsi #2 IsRGB false
Image_T1.vsi #2 LittleEndian false
Image_T1.vsi #2 PixelType uint8
Image_T1.vsi #2 SizeC 3
Image_T1.vsi #2 SizeT 1
Image_T1.vsi #2 SizeX 4206
Image_T1.vsi #2 SizeY 3516
Image_T1.vsi #2 SizeZ 1
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Image_T1.vsi #20 DimensionOrder XYCZT
Image_T1.vsi #20 IsInterleaved true
Image_T1.vsi #20 IsRGB false
Image_T1.vsi #20 LittleEndian false
Image_T1.vsi #20 PixelType uint8
Image_T1.vsi #20 SizeC 3
Image_T1.vsi #20 SizeT 1
Image_T1.vsi #20 SizeX 280
Image_T1.vsi #20 SizeY 163
Image_T1.vsi #20 SizeZ 1
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Image_T1.vsi #3 DimensionOrder XYCZT
Image_T1.vsi #3 IsInterleaved true
Image_T1.vsi #3 IsRGB false
Image_T1.vsi #3 LittleEndian false
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Image_T1.vsi #3 SizeT 1
Image_T1.vsi #3 SizeX 2103
Image_T1.vsi #3 SizeY 1758
Image_T1.vsi #3 SizeZ 1
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Image_T1.vsi #4 DimensionOrder XYCZT
Image_T1.vsi #4 IsInterleaved true
Image_T1.vsi #4 IsRGB false
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Image_T1.vsi #4 SizeX 1052
Image_T1.vsi #4 SizeY 879
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Image_T1.vsi #5 DimensionOrder XYCZT
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Image_T1.vsi #6 DimensionOrder XYCZT
Image_T1.vsi #6 IsInterleaved true
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Image_T1.vsi #8 BitsPerPixel 8
Image_T1.vsi #8 DimensionOrder XYCZT
Image_T1.vsi #8 IsInterleaved true
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Image_T1.vsi #8 SizeT 1
Image_T1.vsi #8 SizeX 3789
Image_T1.vsi #8 SizeY 7582
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Image_T1.vsi #9 BitsPerPixel 8
Image_T1.vsi #9 DimensionOrder XYCZT
Image_T1.vsi #9 IsInterleaved true
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Image_T1.vsi #9 PixelType uint8
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Image_T1.vsi #9 SizeT 1
Image_T1.vsi #9 SizeX 1895
Image_T1.vsi #9 SizeY 3791
Image_T1.vsi #9 SizeZ 1
Series 0 Name label
Series 1 Name Image_T1.vsi #2
Series 10 Name Image_T1.vsi #11
Series 11 Name Image_T1.vsi #12
Series 12 Name 20x
Series 13 Name Image_T1.vsi #14
Series 14 Name Image_T1.vsi #15
Series 15 Name Image_T1.vsi #16
Series 16 Name Image_T1.vsi #17
Series 17 Name Image_T1.vsi #18
Series 18 Name Image_T1.vsi #19
Series 19 Name Image_T1.vsi #20
Series 2 Name Image_T1.vsi #3
Series 20 Name macro image
Series 3 Name Image_T1.vsi #4
Series 4 Name Image_T1.vsi #5
Series 5 Name Image_T1.vsi #6
Series 6 Name overview
Series 7 Name Image_T1.vsi #8
Series 8 Name Image_T1.vsi #9
Series 9 Name Image_T1.vsi #10
label BitsPerPixel 8
label DimensionOrder XYCZT
label IsInterleaved true
label IsRGB false
label LittleEndian false
label PixelType uint8
label SizeC 3
label SizeT 1
label SizeX 8412
label SizeY 7031
label SizeZ 1
macro image BitsPerPixel 8
macro image DimensionOrder XYCZT
macro image IsInterleaved false
macro image IsRGB false
macro image LittleEndian true
macro image PixelType uint8
macro image SizeC 3
macro image SizeT 1
macro image SizeX 190
macro image SizeY 512
macro image SizeZ 1
overview BitsPerPixel 8
overview DimensionOrder XYCZT
overview IsInterleaved true
overview IsRGB false
overview LittleEndian false
overview PixelType uint8
overview SizeC 3
overview SizeT 1
overview SizeX 7578
overview SizeY 15163
overview SizeZ 1
20x Calibration #1 (0.32249193436457624, 0.3224851696897592)
20x Calibration #2 (398.1107327673045, 291.0638972865092)
20x Calibration units #1 10^-6m^1
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20x Creation time (UTC) #1 2017-12-15T15:06:36
20x Creation time (UTC) #2 2017-12-15T15:06:38
20x Creation time (UTC) #3 2017-12-15T15:06:38
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20x External file volume #1 (0, 0, 0, 35800, 20759, 1)
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20x Layer #1 20x
20x Layer #2 20x FocusMap
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20x Microscope Binning (Y) #1 1
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20x Microscope Device Manufacturer #2 Olympus Soft Imaging Solutions
20x Microscope Device Manufacturer #3 Sample Holder
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20x Microscope Device Manufacturer #6 Olympus
20x Microscope Device Manufacturer #7 Olympus
20x Microscope Device Manufacturer #8 Olympus
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20x Microscope Magnification #1 20.0
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20x Microscope Red channel gain #1 2.2270463388939197
20x Microscope Sharpness #1 12
20x Microscope Stage Insert Position #1 (0, 0)
20x Microscope TV Adapter Magnification #1 1.0
20x Microscope Units #1 10^0V^1
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20x Microscope Value #1 9.0
20x Microscope Value #2 0.0
20x Objective Working Distance Units #1 10^-6m^1
20x Objective Working Distance Value #1 600.0000000000001
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Hi Stefanie,

From the information in the error message this appears to be a new issue which we have not seen reported before for VSI.

Do you see this error occurring with all of your images or is it only some specific files? If it is occurring frequently would it be possible to acquire a non confidential sample image from the system which displays the same error?

David

Hi David,

I have just uploaded an image (Image_T30.zip) using the link you previously provided. I hope this helps. Please let me know in case you can't locate the image I uploaded.

Thanks for your help.

Stefanie

Hi Stefanie,

It works fine for me to use Bio-Formats Importer on Fiji or ImageJ to import 4-channel fluorescence .vsi files
acquired by an Olympus microscope. Have you ever updated the Fiji on your system?

Best,
ching

Thank you Stefanie, we received the uploaded sample file and I was able to reproduce the issue as reported and carry out some further testing. This looks like the issue is being caused as Bio-Formats has been unable to determined the type of compression being used on some of the images.

We have previously had 1 other report of VSI files with this compression type but we were unable to determined the compression used. I have updated the existing Trello card with the details from this post https://trello.com/c/fUhH0Cxo/141-cells ... -openbytes.

Do you know what type of compression was used for these files? The current Bio-Formats reader recognises RAW, JPEG, JPEG2000, BMP and PNG.

David Gault

Hi David,

Sorry for not getting back to you earlier. I was in contact with developers from another software that imports VSI files. I got similar issues and they said it is because the files use a rare compression type (lossless JPEG), which is not widely supported. I guess the compression type is indeed the cause of the issues I experience with loading the files. Do you have any idea how I could resolve this?

Thanks very much for your help,

Stefanie

Hi Stefanie,

An alternative you could use the "OlympusImageJPlugin"
https://imagej.net/OlympusImageJPlugin

Follow the installation steps put it to Fiji or ImageJ, then you could load .vsi images by using this plugin. Hopefully, this could solve the problem.

Best,
ching

Hi Stefanie,

Thats good news, thank you for helping us to identify the compression type. We already support using a lossless JPEG codec in other formats so I was able to test this with VSI and the images do appear to be read correctly. As such I have opened a PR to add this codec support to the CellSensReader, which will be reviewed and tested for an upcoming release of Bio-Formats.

https://github.com/openmicroscopy/bioformats/pull/3036

David Gault