> For the complete documentation index, see [llms.txt](https://stereotoolss-organization.gitbook.io/stomics-software-download/llms.txt). Markdown versions of documentation pages are available by appending `.md` to page URLs; this page is available as [Markdown](https://stereotoolss-organization.gitbook.io/stomics-software-download/eco-tools/image-processing.md).

# Image Processing

Image processing and cell segmentation are crucial upstream steps in spatial transcriptomics. Below are the image analysis tools compatible with Stereo-seq imaging workflows, categorized by STOmics Developed Tools (natively integrated with STOmics image formats) and Community Tools (widely used models requiring standard image inputs).&#x20;

{% hint style="info" icon="lightbulb" %}
For performance evaluations and comparative insights across different segmentation models on Stereo-seq datasets, check out our [image benchmarking blog](https://en.stomics.tech/resources/stomics-blog/1162.html).
{% endhint %}

## STOmics Developed

*Tools in this section feature models trained, fine-tuned, or benchmarked directly on Stereo-seq image datasets. They are specifically optimized to handle Stereo-seq data characteristics, such as chip track lines, tissue clarity variations, background noise, etc.*

<table data-card-size="large" data-view="cards"><thead><tr><th></th><th><select multiple><option value="NFb00N6vAlmQ" label="Python Toolkit" color="blue"></option><option value="8gYfIrBVQzd4" label="R Toolkit" color="blue"></option><option value="wACeXfOco5i2" label="CLI" color="blue"></option><option value="U1PDSPCf2AJL" label="Open Source" color="blue"></option><option value="2h0voPCG10K4" label="Commercial" color="blue"></option><option value="Hv2HOwZwRHAG" label="Fine-tuning" color="blue"></option><option value="fzvTuOO6fEEZ" label="Python" color="blue"></option><option value="rh2AUuL4bN4O" label="3D Alignment" color="blue"></option></select></th><th></th><th></th><th data-hidden data-card-cover data-type="image">Cover image</th></tr></thead><tbody><tr><td><h3>cellbin2</h3></td><td><span data-option="NFb00N6vAlmQ">Python Toolkit, </span><span data-option="U1PDSPCf2AJL">Open Source</span></td><td><a href="https://github.com/STOmics/cellbin2" class="button primary medium" data-icon="github">GitHub</a><a href="https://www.biorxiv.org/content/10.1101/2023.02.28.530414v5" class="button secondary medium" data-icon="up-right-from-square">Paper</a></td><td><ul><li><strong>Overview</strong>: A specialized image processing and cell segmentation framework engineered specifically for Stereo-seq tissue images.</li><li><strong>Stereo-seq Optimization</strong>: Trained and benchmarked on native Stereo-seq datasets. Natively handles track-line registration, noise suppression, and varying stain clarity across diverse tissue types.</li><li><strong>Workflow Value</strong>: Directly integrates with <em>SAW</em> pipelines to generate high-precision cell masks and cell-level gene expression matrices (cellbin).</li></ul></td><td data-object-fit="contain"><a href="https://640092348-files.gitbook.io/~/files/v0/b/gitbook-x-prod.appspot.com/o/spaces%2FQfDAr8IqIRwkROXEczGT%2Fuploads%2FMHFH4X1yz503K9pIi7KW%2Fcellbin2.svg?alt=media&amp;token=e3b1434c-a8e7-42e3-930a-29a87699dd3b">cellbin2.svg</a></td></tr><tr><td><h3>CSRefiner</h3></td><td><span data-option="fzvTuOO6fEEZ">Python, </span><span data-option="Hv2HOwZwRHAG">Fine-tuning, </span><span data-option="U1PDSPCf2AJL">Open Source</span></td><td><a href="https://github.com/STOmics/CSRefiner" class="button primary medium" data-icon="github">GitHub</a><a href="https://doi.org/10.1093/bib/bbaf718" class="button secondary medium" data-icon="up-right-from-square">Paper</a></td><td><ul><li><strong>Overview</strong>: A lightweight fine-tuning framework designed to adapt generalist segmentation models (such as <em>Cellpose</em>, <em>StarDist</em>, and <em>cellbin</em>) to specific tissue types using minimal annotated data.</li><li><strong>Stereo-seq Optimization</strong>: Addresses localized segmentation inaccuracies across complex tissue structures; requires as few as 20 image patches for fine-tuning; validated on Stereo-seq DAPI FFPE and H&#x26;E fresh frozen tissue sections.</li><li><strong>Workflow Value</strong>: Produces fine-tuned model weights that integrate seamlessly into <em>SAW</em> or downstream cell segmentation pipelines, directly yielding high-precision cell-level gene expression matrices.</li></ul><p><strong>Blog Post</strong>: <a href="https://en.stomics.tech/resources/stomics-blog/1315.html">CSRefiner: A lightweight framework for fine-tuning cell segmentation models with small datasets</a></p></td><td data-object-fit="contain"><a href="https://640092348-files.gitbook.io/~/files/v0/b/gitbook-x-prod.appspot.com/o/spaces%2FQfDAr8IqIRwkROXEczGT%2Fuploads%2FzHy3W0pqJ2SaetA0R5jJ%2Fcsrefiner.svg?alt=media&amp;token=6775902b-489f-42a0-a528-43ea52b5b838">csrefiner.svg</a></td></tr></tbody></table>

## Community Tools

*Widely used image analysis tools in the bioimage and digital pathology community. These generalist tools can be applied to exported tissue images or bridged into Stereo-seq workflows.*

> *Note: STOmics provides this list for informational purposes only and does not officially support or maintain community-developed tools. We make no guarantees regarding their performance, stability, or compatibility. For technical support, issues, or feature requests, please contact the respective tool developers or consult their official repositories.*

### General Frameworks & Toolkits

*Software platforms and interactive viewers for bioimage visualization, manual annotation, format conversion, and custom workflow development.*

<table data-view="cards"><thead><tr><th></th><th><select multiple><option value="AK5HVENXAdDI" label="Standalone" color="blue"></option><option value="NFb00N6vAlmQ" label="Toolkit" color="blue"></option><option value="bLdmMEBlsxAD" label="Pipeline" color="blue"></option><option value="cGQQQiD2iiWf" label="STOmics Dev" color="blue"></option><option value="5vmJNr4a8VEx" label="Linux" color="blue"></option><option value="h8gbKkC5y7BH" label="Python" color="blue"></option><option value="AefEpxAnTnQE" label="R" color="blue"></option><option value="68zUMQLqQNj5" label="GUI" color="blue"></option><option value="wTgqXhpbvWRi" label="Java" color="blue"></option></select></th><th></th><th></th><th data-hidden data-card-cover data-type="image">Cover image</th></tr></thead><tbody><tr><td><h3>ImageJ/Fiji</h3></td><td><span data-option="wTgqXhpbvWRi">Java, </span><span data-option="68zUMQLqQNj5">GUI</span></td><td><a href="https://github.com/imagej" class="button primary medium" data-icon="github">GitHub-ImageJ</a><a href="https://imagej.net/ij/docs/index.html" class="button primary medium" data-icon="readme">Doc-ImageJ</a><a href="https://github.com/fiji/fiji" class="button primary medium" data-icon="github">GitHub-Fiji</a><a href="https://fiji.sc/" class="button primary medium" data-icon="readme">Doc-Fiji</a></td><td><ul><li><strong>Overview</strong>: An open-source image processing package widely used for analyzing multidimensional biological images.</li><li><strong>How to Bridge</strong>: Excellent for quick visual inspection, manual alignment, and batch image format conversion (e.g., TIFF/PNG) before importing into STOmics tools.</li></ul></td><td data-object-fit="contain"><a href="https://640092348-files.gitbook.io/~/files/v0/b/gitbook-x-prod.appspot.com/o/spaces%2FQfDAr8IqIRwkROXEczGT%2Fuploads%2FOolkno8Efi7roS5Hg3ka%2Fimagej-fiji.svg?alt=media&amp;token=07239f43-e9a8-4ad7-af73-6e58308a955e">imagej-fiji.svg</a></td></tr><tr><td><h3>Napari</h3></td><td><span data-option="h8gbKkC5y7BH">Python, </span><span data-option="68zUMQLqQNj5">GUI</span></td><td><a href="https://github.com/Napari/napari" class="button primary medium" data-icon="github">GitHub</a><a href="https://napari.org/stable/index.html" class="button primary medium" data-icon="readme">Doc</a></td><td><ul><li><strong>Overview</strong>: A fast, interactive 2D/3D image viewer for Python designed for browsing, annotating, and analyzing large multi-dimensional images.</li><li><strong>How to Bridge</strong>: Ideal for Python-native workflows and interactive visual inspection. You can perform manual annotation or segmentation refinement on tissue images, then export cell masks in TIFF or GeoJSON format to integrate seamlessly back into STOmics tools.</li></ul></td><td data-object-fit="contain"><a href="https://640092348-files.gitbook.io/~/files/v0/b/gitbook-x-prod.appspot.com/o/spaces%2FQfDAr8IqIRwkROXEczGT%2Fuploads%2FZd4sqhjJe7dg6r7RbU61%2Fnapari.svg?alt=media&amp;token=a503fa4b-d043-4988-8613-18814c344a28">napari.svg</a></td></tr><tr><td><h3>QuPath</h3></td><td><span data-option="wTgqXhpbvWRi">Java, </span><span data-option="68zUMQLqQNj5">GUI</span></td><td><a href="https://github.com/qupath/qupath" class="button primary medium" data-icon="github">GitHub</a><a href="https://qupath.readthedocs.io/en/stable/" class="button primary medium" data-icon="readme">Doc</a><a href="https://doi.org/10.1038/s41598-017-17204-5" class="button secondary medium" data-icon="up-right-from-square">Paper</a></td><td><ul><li><strong>Overview</strong>: A powerful open-source software for bioimage analysis and digital pathology, optimized for whole-slide image analysis.</li><li><strong>How to Bridge</strong>: Suitable for large Whole Slide Images (WSI). Users can perform tissue annotation or cell segmentation in QuPath and export region masks to STOmics pipelines.</li></ul></td><td data-object-fit="contain"><a href="https://640092348-files.gitbook.io/~/files/v0/b/gitbook-x-prod.appspot.com/o/spaces%2FQfDAr8IqIRwkROXEczGT%2Fuploads%2FZHEgHdZ2F98hQ0f0GJro%2Fqupath.svg?alt=media&amp;token=083c074c-9f97-43e5-9c36-4a14a2df4885">qupath.svg</a></td></tr></tbody></table>

### Cell Segmentation

*Pre-trained deep learning models and algorithmic frameworks for automated single-cell and nuclear segmentation across diverse tissue modalities.*

{% hint style="info" %}
For guidance on using community segmentation outputs to generate cellbin results, check out our [tutorial](https://en.stomics.tech/resources/stomics-blog/1840.html).
{% endhint %}

<table data-view="cards"><thead><tr><th></th><th><select multiple><option value="AK5HVENXAdDI" label="Standalone" color="blue"></option><option value="NFb00N6vAlmQ" label="Toolkit" color="blue"></option><option value="bLdmMEBlsxAD" label="Pipeline" color="blue"></option><option value="cGQQQiD2iiWf" label="STOmics Dev" color="blue"></option><option value="5vmJNr4a8VEx" label="Linux" color="blue"></option><option value="h8gbKkC5y7BH" label="Python" color="blue"></option><option value="AefEpxAnTnQE" label="R" color="blue"></option><option value="68zUMQLqQNj5" label="GUI" color="blue"></option><option value="DmXFXkeDKvNq" label="Deep Learning" color="blue"></option><option value="12z5xOOVucxS" label="Java" color="blue"></option><option value="x4cNOIipycab" label="CLI" color="blue"></option></select></th><th></th><th></th><th data-hidden data-card-cover data-type="image">Cover image</th></tr></thead><tbody><tr><td><h3>Cellpose</h3></td><td><span data-option="h8gbKkC5y7BH">Python, </span><span data-option="68zUMQLqQNj5">GUI, </span><span data-option="DmXFXkeDKvNq">Deep Learning</span></td><td><a href="https://github.com/MouseLand/cellpose" class="button primary medium" data-icon="github">GitHub</a><a href="https://cellpose.readthedocs.io/en/latest/" class="button primary medium" data-icon="readme">Doc</a></td><td><ul><li><strong>Overview</strong>: A generalist deep learning algorithm for cell segmentation across diverse cell shapes, sizes, and cellular modalities.</li><li><strong>How to Bridge</strong>: Performs single-cell or nuclear segmentation on multi-stained tissue images via its GUI or Python API, then exports standard cell masks in TIFF or GeoJSON format to integrate into STOmics analysis tools for high-precision cell binning.</li></ul></td><td data-object-fit="contain"><a href="https://640092348-files.gitbook.io/~/files/v0/b/gitbook-x-prod.appspot.com/o/spaces%2FQfDAr8IqIRwkROXEczGT%2Fuploads%2Fu7Cuw4zAiooSnDUCQ3gb%2Fcellpose.svg?alt=media&amp;token=f8f378dc-d7df-4437-af55-6e5b22e8cb2a">cellpose.svg</a></td></tr><tr><td><h3>StarDist</h3></td><td><span data-option="h8gbKkC5y7BH">Python, </span><span data-option="12z5xOOVucxS">Java, </span><span data-option="DmXFXkeDKvNq">Deep Learning</span></td><td><a href="https://github.com/stardist/stardist" class="button primary medium" data-icon="github">GitHub</a><a href="https://stardist.net/" class="button primary medium" data-icon="readme">Doc</a></td><td><ul><li><strong>Overview</strong>: A star-convex object detection model optimized for dense, star-convex shapes like cell nuclei.</li><li><strong>How to Bridge</strong>: Ideal for nuclear-stained (e.g., DAPI) raw tissue images. Output nuclear masks exported in TIFF or GeoJSON format can be imported into STOmics analysis tools to assist in nuclear-guided cell binning.</li></ul></td><td data-object-fit="contain"><a href="https://640092348-files.gitbook.io/~/files/v0/b/gitbook-x-prod.appspot.com/o/spaces%2FQfDAr8IqIRwkROXEczGT%2Fuploads%2FHY6gefDW0hTPBX6iyxqT%2Fstardist.svg?alt=media&amp;token=1e62d990-d3e3-4fa1-89f2-de5e33323113">stardist.svg</a></td></tr><tr><td><h3>Omnipose</h3></td><td><span data-option="h8gbKkC5y7BH">Python, </span><span data-option="DmXFXkeDKvNq">Deep Learning</span></td><td><a href="https://github.com/kevinjohncutler/omnipose" class="button primary medium" data-icon="github">GitHub</a><a href="https://omnipose.readthedocs.io/" class="button primary medium" data-icon="readme">Doc</a></td><td><ul><li><strong>Overview</strong>: A deep learning-based segmentation algorithm designed for extremely varied cell shapes, bacterial microscopy, and dense tissues.</li><li><strong>How to Bridge</strong>: Suitable for non-standard or highly complex cell morphology datasets where generalist models like Cellpose fall short. Output nuclear masks can be directly imported into STOmics analysis tools.</li></ul></td><td data-object-fit="contain"><a href="https://640092348-files.gitbook.io/~/files/v0/b/gitbook-x-prod.appspot.com/o/spaces%2FQfDAr8IqIRwkROXEczGT%2Fuploads%2FjS3wzGCj3Y9XTtdm7NR9%2Fomnipose.svg?alt=media&amp;token=f23b3777-8286-4bb0-864e-e6f2bc844d26">omnipose.svg</a></td></tr><tr><td><h3>DeepCell</h3></td><td><span data-option="h8gbKkC5y7BH">Python, </span><span data-option="x4cNOIipycab">CLI, </span><span data-option="DmXFXkeDKvNq">Deep Learning</span></td><td><a href="https://github.com/vanvalenlab/deepcell-tf" class="button primary medium" data-icon="github">GitHub</a><a href="https://deepcell.readthedocs.io/en/master/" class="button primary medium" data-icon="readme">Doc</a></td><td><ul><li><strong>Overview</strong>: A deep learning library featuring pre-trained models (like Mesmer) for cell segmentation in tissue imaging.</li><li><strong>How to Bridge</strong>: Provides robust segmentation across diverse tissue modalities. Output nuclear or membrane masks can be converted to standard cell segmentation formats for STOmics tools ingestion.</li></ul></td><td data-object-fit="contain"><a href="https://640092348-files.gitbook.io/~/files/v0/b/gitbook-x-prod.appspot.com/o/spaces%2FQfDAr8IqIRwkROXEczGT%2Fuploads%2FzJjiWJynWHFxxTwBrAuQ%2Fdeepcell.svg?alt=media&amp;token=7ab1fc00-c638-473f-a88b-0f2b5dbb7e8c">deepcell.svg</a></td></tr></tbody></table>

### Image Registration

*Specialized tools and plugins designed for multi-channel image stitching and multi-slice registration.*

<table data-view="cards"><thead><tr><th></th><th><select multiple><option value="NFb00N6vAlmQ" label="Toolkit" color="blue"></option><option value="5vmJNr4a8VEx" label="Linux" color="blue"></option><option value="h8gbKkC5y7BH" label="Python" color="blue"></option><option value="AefEpxAnTnQE" label="R" color="blue"></option><option value="68zUMQLqQNj5" label="GUI" color="blue"></option><option value="wTgqXhpbvWRi" label="Java" color="blue"></option><option value="U1xffhzrRYhG" label="ImageJ Plugin" color="blue"></option><option value="gVKiiK5jzAu2" label="3D Alignment" color="blue"></option></select></th><th></th><th></th><th data-hidden data-card-cover data-type="image">Cover image</th></tr></thead><tbody><tr><td><h3>TrakEM2 (ImageJ Plugin)</h3></td><td><span data-option="wTgqXhpbvWRi">Java, </span><span data-option="U1xffhzrRYhG">ImageJ Plugin, </span><span data-option="68zUMQLqQNj5">GUI</span></td><td><a href="https://github.com/trakem2/TrakEM2" class="button primary medium" data-icon="github">GitHub</a><a href="https://imagej.net/plugins/trakem2/" class="button primary medium" data-icon="readme">Doc</a></td><td><ul><li><strong>Overview</strong>: An ImageJ plugin for morphological data mining, three-dimensional modeling and image stitching, registration, editing and annotation.</li><li><strong>How to Bridge</strong>: Can be used to manually or semi-automatically align multi-slice or multi-channel tissue images prior to downstream spatial analysis.</li></ul></td><td data-object-fit="contain"><a href="https://640092348-files.gitbook.io/~/files/v0/b/gitbook-x-prod.appspot.com/o/spaces%2FQfDAr8IqIRwkROXEczGT%2Fuploads%2FoqfRKcvztJDoBKC3SRpy%2Ftrakem2.svg?alt=media&amp;token=c85d5282-f6a3-4d14-a9f6-4bfebba30b26">trakem2.svg</a></td></tr></tbody></table>
