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Introduction to the ImageJ Ecosystem

Edward L. Evans III, Ph.D

Eliceiri Lab / LOCI

elevans2@wisc.edu

Slides at

imagej.net/events/presentations

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Outline

What is ImageJ?

Understanding the ImageJ/Fiji family

What can ImageJ/Fiji do?

Plugins: Biovoxxel, StarDist etc…
Scripting and automation with macros and scripts

Learning more about ImageJ/Fiji

Online resources

ImageJ on other platforms

PyImageJ
KNIME
JIPipe

Additional resources
Bonus: Demos!

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What is ImageJ?

Primarily* a desktop application for scientific image analysis.
Reusable set of libraries and shared framework for image analysis
Open source with strong community support.
Written in Java, enabling ImageJ to run in diverse computing environments.
Multiple “flavors” of ImageJ:

* ImageJ2’s headless feature allows ImageJ2 to run on distributed computing networks

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What is ImageJ?

Stable version of the original ImageJ (1.x) has been continuously developed since 1997.

Originally developed by Wayne Rasband in 1997 as a cross-platform version of “NIH Image”.

ImageJ2 includes the latest version of ImageJ (1.x) with a legacy layer for backwards compatibility.

The legacy layer enables transparent conversions between ImageJ and ImageJ2 data structures as needed.

Fiji is a “batteries included” distribution of ImageJ2.

Wayne Rasband

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What is Fiji?

Fiji Is Just ImageJ

Fiji is a “batteries included” distribution of ImageJ2.
Advantages for end users:

Easy to install (Linux, MacOS, Windows)
Automatic updates
Bundles a lot of plugins (and more via update sites!)
Comprehensive documentation
Visit https://imagej.net/software/imagej2/ for more details

Advantages for developers:

Open source project (GitHub)
Expanded language support (Groovy, Python, JavaScript, . etc…) eases the development of scripts and plugins

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What can Fiji do?

Stitching

Big Data

Segmentation

Tracking

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Plugins

What is a plugin?

Plugins are special-purpose software components that extend the functionality of the ImageJ ecosystem.

There are four tiers of plugins:

Core ImageJ plugins (1000+)
Core Fiji plugins (1000+)
Plugins installable from Update Sites.
Additional manually-installed plugins available from various online sources.

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Fiji Plugin: General image processing with Biovoxxel

BioVoxxel Toolbox

Extended Particle Analyzer

https://imagej.net/BioVoxxel_Toolbox#Extended_Particle_Analyzer

Threshold Check

https://imagej.net/BioVoxxel_Toolbox#Threshold_Check

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Fiji Plugin: MorphoLibJ

Features:

Morphological filtering for 2D/3D and binary or grey level images: erosion & dilation, closing & opening, morphological gradient & Laplacian, top-hat, etc…
Morphological reconstruction for 2D/3D and binary or grey level images, allowing fast detection of regional or extended extrema, border removal, hole filling, attribute filtering, etc…
Watershed segmentation + GUI, making it possible to segment 2D/3D images.
2D/3D measurements: photometric (intensity) and morphometric measurements such as volume, surface area, inertia ellipse/ellipsoid, etc…
Binary / label images utilities for removing or keeping largest connected component, perform size opening, fill holes, remove borders, etc…

https://imagej.net/plugins/morphological-segmentation

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Fiji Plugin: TrackMate

Features:

Single particle tracking
Simple/intuitive interface
Segmentation, filtering, particle-linking process visualized in 2D or 3D.

https://imagej.net/plugins/trackmate/

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Fiji Plugin: Trainable Weka Segmentation

Features:

A Weka machine learning based plugin that produces pixel-based segmentation results.
Can segment 2D and 3D data.

https://imagej.net/plugins/tws/index

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Fiji Plugin: StarDist

Features:

Detects cell/nuclei with star-convex shape priors (i.e. “circular” objects)
Can be used to apply pre-trained models to new 2D/2D+time images.
Full featured Python package for training new models and 3D data

https://imagej.net/plugins/stardist

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Fiji Plugin: Coloc 2

Features:

Pixel intensity correlation over space methods of Pearson, Manders, Costes, Li and more
Scatterplots, analysis, automatic thresholding and statistical significance testing
Supports ROIs.

https://imagej.net/plugins/coloc-2

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Registration...

Feature Extraction

https://imagej.net/Feature_Extraction

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Registration...

https://imagej.net/Register_Virtual_Stack_Slices

Feature Extraction

https://imagej.net/Feature_Extraction

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Registration...

TrakEM2

https://imagej.net/TrakEM2

https://imagej.net/Register_Virtual_Stack_Slices

Feature Extraction

https://imagej.net/Feature_Extraction

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Registration...

TrakEM2

https://imagej.net/TrakEM2

Feature Extraction

https://imagej.net/Feature_Extraction

bUnwarpJ

https://imagej.net/BUnwarpJ

https://imagej.net/Register_Virtual_Stack_Slices

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Scripting and automation

Why use scripts?

Reproducible science
Document your work
Automate your analysis
Share with the world

Supported Languages:

Groovy, ImageJ Macro, Python(Jython), JavaScript, Ruby(JRuby), Lisp(Clojure), R(Renjin), Java, Matlab, BeanShell, Scala

https://imagej.net/scripting/script-editor

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Scripting and automation

ImageJ Macro language:

Less powerful
Easy to learn and use

ImageJ Macro Recorder

https://imagej.net/scripting/macro#the-recorder

Built-in Macro Functions list

https://imagej.net/ij/developer/macro/functions.html

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Where to learn more?

https://imagej.net

http://imagej.org

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Search on imagej.net to learn more about your topic

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The original ImageJ documentation at imagej.org

Original ImageJ user guide
Extensive documentation on commands and macros
Tutorials, examples, image processing guides

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Learn and teach others on the image.sc forum

https://forum.image.sc/

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ImageJ and more on the forum!

There is more than just ImageJ help on the forum!
Have a basic image processing question? Ask the forum!

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ImageJ on other platforms

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PyImageJ: A library for integrating ImageJ and Python

PyImageJ:

A Python based package that integrates ImageJ into the Python ecosystem.
Use ImageJ (and plugins) with Python packages like: NumPy, SciPy, scikit-image, CellProfiler, OpenCV, ITK and more
Full support for existing macros and scripts in all supported languages.
Headless and interactive modes.

https://pyimagej.readthedocs.io/en/latest/01-Starting-PyImageJ.html

Rueden et al. Nature Methods 2022

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ImageJ integration in Konstanz Information Miner (KNIME)

Java-based program
Robust image analysis operations
Supports macros, scripts and plugins

Java-based program
Intuitive visual workflow model for programing
Powerful data organization tools

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Example workflow: Nuclear Ring Segmentation, Analysis and Tracking (NR-SAT)

Connect nodes to flow data between ImageJ functions

Intuitive user interface

Powerful data frame tools (similar to pandas)

Transpose
Join
Concatenate

Macro and script support

Use your existing macros and scripts

Live data interrogation

Examine your data at any point in the workflow

Discover and share workflows on the KNIME Hub

https://hub.knime.com/

Evans et al. Viruses 2022

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Example workflow: Nuclear Ring Segmentation, Analysis and Tracking (NR-SAT)

Evans et al. Viruses 2022

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Example workflow: Nuclear Ring Segmentation, Analysis and Tracking (NR-SAT)

Evans et al. Viruses 2022

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Example workflow: Nuclear Ring Segmentation, Analysis and Tracking (NR-SAT)

Evans et al. Viruses 2022

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Example workflow: Nuclear Ring Segmentation, Analysis and Tracking (NR-SAT)

Evans et al. Viruses 2022

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Example workflow: Nuclear Ring Segmentation, Analysis and Tracking (NR-SAT)

Evans et al. Viruses 2022

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JIPipe: visual batch processing for ImageJ

Create ImageJ macros without any programming
“Flowchart” data flow scheme
Extensive documentation

Example and tutorials
Developer documentation

https://www.jipipe.org/

Gerst et al. Nature Methods 2023

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Center for Open Bioimage Analysis

openbioimageanalysis.org/

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COBA: Driving Biological Projects

Check out what we do!
Download and use macros, scripts and workflows
Collaborate with us!

openbioimageanalysis.org/

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BioImaging North America (BINA)

6 Working Groups: Communications, Corporate Partners, Diversity, Equity & Inclusion, Image Informatics, Quality Control & Data Management, Training & Education

www.BioImagingNorthAmerica.org/join

contact@bioimagingna.org

@BioimagingNA

Events and Newsletter each month!

Mission: Engaging bioimaging scientists across North America by creating an inclusive and supportive community to share, advance and succeed together.