SEM image analysis: DiameterJ with Auto thresholding #1970
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| name: sem_analysis | ||
| owner: bgruening | ||
| description: Scanning electron micrographs (SEM) fiber analysis using FIJI, DiameterJ and Auto-thresholding. | ||
| long_description: | | ||
| The SEM Analysis tool segments fibers and measures their diameters, and pores with FIJI, DiameterJ and Auto-thresholding | ||
| remote_repository_url: https://github.com/bgruening/galaxytools/tree/master/tools/sem_analysis | ||
| homepage_url: https://github.com/bgruening/galaxytools/tree/master/tools/sem_analysis | ||
| type: | ||
| categories: | ||
| - Imaging | ||
| maintainers: | ||
| anuprulez |
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| #!/usr/bin/env python3 | ||
| """Convert all DiameterJ radius-plot TIFFs in a result directory to PNG.""" | ||
| import sys | ||
| from pathlib import Path | ||
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| from PIL import Image | ||
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| def main() -> None: | ||
| result_dir = Path(sys.argv[1]) | ||
| for source in result_dir.glob("*_Radius Plot.tif"): | ||
| destination = source.with_suffix(".png") | ||
| with Image.open(source) as image: | ||
| image.save(destination) | ||
| source.unlink() | ||
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| if __name__ == "__main__": | ||
| main() | ||
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| <tool id="sem_analysis" name="Scanning electron micrographs (SEM) fiber analysis" version="@TOOL_VERSION@+galaxy@VERSION_SUFFIX@" profile="@PROFILE@"> | ||
| <description>segment fibers and measure their diameters</description> | ||
| <macros> | ||
| <token name="@TOOL_VERSION@">1-018</token> | ||
| <token name="@VERSION_SUFFIX@">0</token> | ||
| <token name="@PROFILE@">25.0</token> | ||
| </macros> | ||
| <creator> | ||
| <person givenName="Anup" familyName="Kumar" email="kumara@informatik.uni-freiburg.de"/> | ||
| <organization name="European Galaxy Team" url="https://galaxyproject.org/eu/"/> | ||
| </creator> | ||
| <requirements> | ||
| <container type="docker">quay.io/galaxy/sem-analysis-fiji-diameterj:@TOOL_VERSION@</container> | ||
| </requirements> | ||
| <required_files> | ||
| <include path="convert_radius_plots.py"/> | ||
| </required_files> | ||
| <command detect_errors="exit_code"><![CDATA[ | ||
| mkdir -p results && | ||
| export ANALYSIS_RESULTS_DIR='results' && | ||
| ln -s '$input_image' input.tif && | ||
| srm_args='' | ||
| #if $segmentation.workflow in ('recursive-srm', 'srm-auto-thresholding'): | ||
| srm_args='--srm-q $segmentation.srm_q' | ||
| #end if | ||
| #if $calibration.source == 'pixel_size': | ||
| calibration_args='--pixel-size-um $calibration.pixel_size_um' | ||
| #else: | ||
| calibration_args='--hfw-um $calibration.hfw_um' | ||
| #end if | ||
| mode_args='' | ||
| #if $analysis_mode == 'qc': | ||
| mode_args='--skip-diameterj' | ||
| #end if | ||
| #if $analysis_mode == 'diameterj': | ||
| /usr/local/bin/sem-analysis --input input.tif --output results --segmentation '$segmentation.workflow' --threshold-methods '#echo ",".join($threshold_methods)#' \$srm_args \$calibration_args \$mode_args --crop-bottom '$advanced.crop_bottom' && | ||
| python3 '$__tool_directory__/convert_radius_plots.py' results | ||
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| #else: | ||
| /usr/local/bin/sem-analysis --input input.tif --output results --segmentation '$segmentation.workflow' --threshold-methods '#echo ",".join($threshold_methods)#' \$srm_args \$calibration_args \$mode_args --crop-bottom '$advanced.crop_bottom' | ||
| #end if | ||
| ]]></command> | ||
| <inputs> | ||
| <param name="input_image" type="data" format="tiff" label="Scanning electron micrographs" help="A 2-D TIFF image. RGB TIFFs are converted to grayscale."/> | ||
| <conditional name="segmentation"> | ||
| <param name="workflow" type="select" label="Segmentation workflow"> | ||
| <option value="auto-thresholding" selected="true">Auto Thresholding (Traditional)</option> | ||
| <option value="recursive-srm">Recursive SRM with Auto Thresholding (SRM)</option> | ||
| <option value="srm-auto-thresholding">SRM with Auto Thresholding (Mixed)</option> | ||
| </param> | ||
| <when value="auto-thresholding"/> | ||
| <when value="recursive-srm"> | ||
| <param name="srm_q" type="integer" value="100" min="1" label="Starting SRM granularity (q)"/> | ||
|
Owner
There was a problem hiding this comment. use argument here and elsewhere? |
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| </when> | ||
| <when value="srm-auto-thresholding"> | ||
| <param name="srm_q" type="integer" value="100" min="1" label="SRM granularity (q)"/> | ||
| </when> | ||
| </conditional> | ||
| <param name="threshold_methods" type="select" multiple="true" optional="false" label="Thresholding methods" help="Select one or more Fiji Auto Threshold methods. Each selected method produces a separate segmentation candidate."> | ||
| <option value="default" selected="true">Default</option> | ||
| <option value="huang">Huang</option> | ||
| <option value="huang2">Huang2</option> | ||
| <option value="intermodes">Intermodes</option> | ||
| <option value="isodata">IsoData</option> | ||
| <option value="li">Li</option> | ||
| <option value="maxentropy">MaxEntropy</option> | ||
| <option value="mean">Mean</option> | ||
| <option value="minerror">MinError(I)</option> | ||
| <option value="minimum">Minimum</option> | ||
| <option value="moments">Moments</option> | ||
| <option value="otsu">Otsu</option> | ||
| <option value="percentile">Percentile</option> | ||
| <option value="renyientropy">RenyiEntropy</option> | ||
| <option value="shanbhag">Shanbhag</option> | ||
| <option value="triangle">Triangle</option> | ||
| <option value="yen">Yen</option> | ||
| </param> | ||
| <param name="analysis_mode" type="select" label="Analysis mode"> | ||
| <option value="diameterj" selected="true">Full DiameterJ analysis</option> | ||
| <option value="qc">Segmentation and QC only</option> | ||
| </param> | ||
| <conditional name="calibration"> | ||
| <param name="source" type="select" label="Pixel calibration"> | ||
| <option value="hfw" selected="true">Derive from horizontal field width</option> | ||
| <option value="pixel_size">Specify pixel size directly</option> | ||
| </param> | ||
| <when value="hfw"> | ||
| <param name="hfw_um" type="float" value="27.04" min="0.01" max="1000" label="Horizontal field width (µm)" help="Physical width of the full image, as recorded by the SEM. The default 27.04 µm corresponds to 0.02640625 µm/pixel for a 1024-pixel-wide image; replace it with the value from your image metadata. Increasing or decreasing HFW scales all derived physical sizes by the same proportion, but does not change pixel-based segmentation."/> | ||
| </when> | ||
| <when value="pixel_size"> | ||
| <param name="pixel_size_um" type="float" value="0.02640625" min="0.000001" max="1000" label="Pixel size (µm/pixel)" help="Physical size represented by one image pixel, as recorded by the SEM or calculated from a known scale. The default 0.02640625 µm/pixel corresponds to a 27.04 µm horizontal field width for a 1024-pixel-wide image; replace it with the value from your image metadata. Increasing or decreasing pixel size scales all derived physical sizes by the same proportion, but does not change pixel-based segmentation."/> | ||
| </when> | ||
| </conditional> | ||
| <section name="advanced" title="Segmentation options" expanded="false"> | ||
| <param name="crop_bottom" type="integer" value="59" min="0" label="Crop from bottom (pixels)" help="Removes the SEM instrument footer before analysis."/> | ||
| </section> | ||
| </inputs> | ||
| <outputs> | ||
| <collection name="results_collection" type="list" label="${tool.name}: results for ${on_string}"> | ||
| <discover_datasets pattern="__name_and_ext__" directory="results"/> | ||
| </collection> | ||
| </outputs> | ||
| <tests> | ||
| <test expect_num_outputs="1"> | ||
| <param name="input_image" value="PVA-A_004.tif"/> | ||
| <conditional name="segmentation"> | ||
| <param name="workflow" value="auto-thresholding"/> | ||
| </conditional> | ||
| <param name="analysis_mode" value="qc"/> | ||
| <param name="threshold_methods" value="default,otsu"/> | ||
| <output_collection name="results_collection" type="list" count="21"/> | ||
| </test> | ||
| <test expect_num_outputs="1"> | ||
| <param name="input_image" value="PVA-A_004.tif"/> | ||
| <conditional name="segmentation"> | ||
| <param name="workflow" value="auto-thresholding"/> | ||
| </conditional> | ||
| <param name="analysis_mode" value="qc"/> | ||
| <output_collection name="results_collection" type="list" count="12"/> | ||
| </test> | ||
| <test expect_num_outputs="1"> | ||
| <param name="input_image" value="PVA-A_004.tif"/> | ||
| <conditional name="segmentation"> | ||
| <param name="workflow" value="srm-auto-thresholding"/> | ||
| <param name="srm_q" value="80"/> | ||
| </conditional> | ||
| <param name="analysis_mode" value="qc"/> | ||
| <output_collection name="results_collection" type="list" count="12"/> | ||
| </test> | ||
| <test expect_num_outputs="1"> | ||
| <param name="input_image" value="PVA-A_004.tif"/> | ||
| <conditional name="segmentation"> | ||
| <param name="workflow" value="recursive-srm"/> | ||
| <param name="srm_q" value="100"/> | ||
| </conditional> | ||
| <param name="analysis_mode" value="qc"/> | ||
| <output_collection name="results_collection" type="list" count="12"/> | ||
| </test> | ||
| <test expect_num_outputs="1"> | ||
| <param name="input_image" value="PVA-A_004.tif"/> | ||
| <conditional name="segmentation"> | ||
| <param name="workflow" value="auto-thresholding"/> | ||
| </conditional> | ||
| <param name="threshold_methods" value="huang"/> | ||
| <param name="analysis_mode" value="qc"/> | ||
| <output_collection name="results_collection" type="list" count="12"/> | ||
| </test> | ||
| <test expect_num_outputs="1"> | ||
| <param name="input_image" value="PVA-A_004.tif"/> | ||
| <conditional name="segmentation"> | ||
| <param name="workflow" value="auto-thresholding"/> | ||
| </conditional> | ||
| <param name="threshold_methods" value="huang2,intermodes,isodata"/> | ||
| <param name="analysis_mode" value="qc"/> | ||
| <conditional name="calibration"> | ||
| <param name="source" value="pixel_size"/> | ||
| <param name="pixel_size_um" value="0.03"/> | ||
| </conditional> | ||
| <output_collection name="results_collection" type="list" count="30"/> | ||
| </test> | ||
| <test expect_num_outputs="1"> | ||
| <param name="input_image" value="PVA-A_004.tif"/> | ||
| <conditional name="segmentation"> | ||
| <param name="workflow" value="srm-auto-thresholding"/> | ||
| <param name="srm_q" value="60"/> | ||
| </conditional> | ||
| <param name="threshold_methods" value="li"/> | ||
| <param name="analysis_mode" value="qc"/> | ||
| <output_collection name="results_collection" type="list" count="12"/> | ||
| </test> | ||
| <test expect_num_outputs="1"> | ||
| <param name="input_image" value="PVA-A_004.tif"/> | ||
| <conditional name="segmentation"> | ||
| <param name="workflow" value="srm-auto-thresholding"/> | ||
| <param name="srm_q" value="120"/> | ||
| </conditional> | ||
| <param name="threshold_methods" value="maxentropy,mean,minerror"/> | ||
| <param name="analysis_mode" value="qc"/> | ||
| <output_collection name="results_collection" type="list" count="30"/> | ||
| </test> | ||
| <test expect_num_outputs="1"> | ||
| <param name="input_image" value="PVA-A_004.tif"/> | ||
| <conditional name="segmentation"> | ||
| <param name="workflow" value="recursive-srm"/> | ||
| <param name="srm_q" value="64"/> | ||
| </conditional> | ||
| <param name="threshold_methods" value="minerror"/> | ||
| <param name="analysis_mode" value="qc"/> | ||
| <output_collection name="results_collection" type="list" count="12"/> | ||
| </test> | ||
| <test expect_num_outputs="1"> | ||
| <param name="input_image" value="PVA-A_004.tif"/> | ||
| <conditional name="segmentation"> | ||
| <param name="workflow" value="recursive-srm"/> | ||
| <param name="srm_q" value="80"/> | ||
| </conditional> | ||
| <param name="threshold_methods" value="moments,percentile,renyientropy,shanbhag,triangle,yen"/> | ||
| <param name="analysis_mode" value="qc"/> | ||
| <conditional name="calibration"> | ||
| <param name="source" value="hfw"/> | ||
| <param name="hfw_um" value="30"/> | ||
| </conditional> | ||
| <output_collection name="results_collection" type="list" count="57"/> | ||
| </test> | ||
| </tests> | ||
| <help>< | ||
| methods directly to the cropped image. The available methods are Default, | ||
| Huang, Huang2, Intermodes, | ||
| IsoData, Li, MaxEntropy, Mean, MinError(I), Minimum, Moments, Otsu, | ||
| Percentile, RenyiEntropy, Shanbhag, Triangle, and Yen. | ||
| * **SRM with Auto Thresholding** applies one [Statistical Region Merging](https://imagej.net/plugins/statistical-region-merging) pass at | ||
| the selected ``q``, converts the result to 8-bit, and applies the selected | ||
| Auto Threshold methods. | ||
| * **Recursive SRM** runs exactly one recursive workflow. It successively applies | ||
| SRM at ``q``, ``q/2``, ``q/4``, and ``q/8`` (with a minimum of 1), then | ||
| applies the selected threshold methods to the final SRM image. | ||
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| Use **Thresholding methods** to choose one or more of these methods. The same | ||
| selection is applied after SRM for either SRM-based workflow. All 17 methods are | ||
| selected by default. | ||
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| **Analysis mode** | ||
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| * **Full DiameterJ analysis** creates the selected segmentation masks and QC | ||
| results, then runs DiameterJ 1.018 on every mask. Candidates are not rejected | ||
| based on their black-pixel fraction, so review the segmentation and overlay | ||
| montages when a threshold method produces an implausible mask. | ||
| * **Segmentation and QC only** skips DiameterJ analysis. It is faster and | ||
| is useful for comparing candidate masks before a full run. | ||
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| **Calibration and interpretation** | ||
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| QC measurements are reported in pixels and micrometres. Choose one of the | ||
| following calibration sources: | ||
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| * **Horizontal field width** is the physical width represented by the entire | ||
| uncropped image. Pixel size is calculated as ``HFW / image width in pixels``. | ||
| The default is 27.04 µm and should be replaced with the value recorded by the | ||
| microscope for the input image. | ||
| * **Pixel size** directly specifies micrometres per pixel. Use this when the | ||
| calibrated pixel spacing is already known. | ||
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| Cropping the bottom does not change image width, so it does not alter an | ||
| HFW-derived pixel size. DiameterJ runs in its validated native-pixel mode because | ||
| physical calibration can change its legacy particle-size filtering. Use the | ||
| ``pixel_size_um`` field in ``qc_summary.csv`` when converting DiameterJ | ||
| measurements to physical units. | ||
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| **Segmentation options** | ||
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| * **Crop from bottom** removes a fixed number of pixel rows before analysis. The | ||
| default of 59 is intended for an SEM annotation or instrument footer. Set it | ||
| to 0 when the image contains no footer. It must be smaller than image height. | ||
| * **SRM granularity (q)** controls region merging in SRM with Auto Thresholding | ||
| and Recursive SRM modes. It must be positive. Larger values generally | ||
| preserve more, smaller regions; lower values generally merge regions more | ||
| aggressively. The default is 100. | ||
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| **Outputs** | ||
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| All generated files are returned as one list collection in the Galaxy history. | ||
| Each workflow produces two PNG review sheets: a segmentation montage containing | ||
| the cropped original and every binary mask, and an overlay montage containing | ||
| the original and every boundary overlay. It also produces each candidate as a | ||
| separate, display-ready PNG and as a matching TIFF used for DiameterJ analysis. | ||
| Each workflow produces one segmented PNG and one matching TIFF per selected | ||
| threshold method. | ||
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| Dataset names retain the input stem, segmentation family, SRM settings where | ||
| applicable, and threshold method. For each mask, QC-only mode also includes a | ||
| boundary-overlay PNG and diameter-samples CSV; the collection contains one | ||
| QC summary CSV. Full mode adds the DiameterJ total summary, comparison | ||
| image, pore data, radius histogram, characteristic lengths, and radius plot for | ||
| each mask. Every DiameterJ radius-plot TIFF is converted to PNG before Galaxy | ||
| discovers the collection. The resulting collection size depends on the number of | ||
| selected threshold methods and the analysis mode. | ||
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| The QC diameter samples are distance-transform measurements on skeleton pixels. | ||
| They are intended as an independent quality check; DiameterJ output is the | ||
| primary result in full-analysis mode. | ||
| ]]></help> | ||
| <citations> | ||
| <citation type="doi">10.1016/j.biomaterials.2015.05.015</citation> | ||
| </citations> | ||
| </tool> | ||
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why is this actually needed? Is TIFF not useful?
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I agree. But not sure what our collaborators will require. We share them the tool and fix other feedbacks along with this.