Overview

Flow cytometry calibration is often distributed across several disconnected tools: instrument software, notebooks, scripts, spreadsheets, manually selected peaks, copied constants, and exported configuration files. This fragmentation makes calibration procedures difficult to audit, reproduce, compare, and reuse.

RosettaX brings these steps into a structured application. A calibration session can be configured, inspected, fitted, saved, and reused without separating the scientific model from the practical workflow. This is particularly useful when calibration results need to remain traceable across instruments, detector settings, reference materials, and analysis pipelines.

RosettaX currently focuses on two main calibration workflows:

• fluorescence calibration

• scattering calibration

The fluorescence workflow supports calibration from reference intensity data. The scattering workflow connects measured cytometry signals with optical parameters such as particle size, refractive index, detector configuration, and scattering intensity.

Features

RosettaX currently provides:

• fluorescence calibration workflows

• scattering calibration workflows

• FCS data loading

• interactive histogram inspection

• peak identification and peak finding

• calibration fitting

• reusable calibration export

• runtime profile management

• detector and optical parameter presets

• support for default and custom parameter configurations

• local graphical application behavior suitable for repeated calibration work

Why RosettaX

RosettaX is intended to reduce the amount of manual and implicit work involved in flow cytometry calibration.

Typical calibration workflows often require users to move between acquisition software, custom scripts, spreadsheets, and manually recorded parameters. This can make it difficult to know exactly which data, peaks, settings, assumptions, and fitted models produced a given calibration result.

RosettaX addresses this by keeping the workflow organized around explicit steps:

• load the relevant cytometry data

• select or define the calibration context

• inspect the signal distributions

• identify the calibration peaks

• fit the calibration relation

• review the result

• export a reusable calibration file

This makes the calibration process easier to document, compare, repeat, and adapt to new experiments.

Typical Workflow

A typical RosettaX session follows these steps:

1. launch the local application

2. load or select a runtime profile

3. import the relevant FCS data

4. choose the fluorescence or scattering calibration workflow

5. configure the detector, optical, and calibration parameters

6. inspect histograms and identify calibration peaks

7. fit the calibration model

8. review the calibration result

9. save the calibration for later reuse

The exact procedure depends on the selected workflow, but the application is organized so that each step remains visible and traceable.

On smaller screens, open Menu to access navigation and saved profiles. Graphs use a viewport-aware default height; an explicit graph height in your profile remains in effect. Calibration reference tables keep headers visible while scrolling and show cell-specific feedback for invalid numeric entries. Blank cells remain available for completing a table in stages.

Scientific Scope

RosettaX is intended for research and development workflows involving flow cytometry calibration. It focuses on calibration analysis and calibration file generation rather than instrument control.

The application is especially relevant when experimental cytometry data, reference materials, optical parameters, detector settings, and reusable analysis profiles need to be managed together in a structured workflow.

For scattering calibration, RosettaX is designed to connect measured cytometry signals with optical modeling concepts, including particle size, refractive index, collection geometry, detector configuration, and scattering intensity. This makes it suitable for workflows where optical interpretation and instrument calibration need to remain linked.

Data Handling

RosettaX runs locally.

The application does not upload FCS files, calibration profiles, calibration results, or generated configuration files to a remote service. Data remain on the machine running the software unless the user explicitly moves, uploads, or shares them outside the application.

Project Status

RosettaX is currently in alpha development.

The application is usable for development, testing, and iterative calibration workflow design. However, the public API, internal organization, and graphical workflows may still change. New users should expect active development and occasional breaking changes.

Current development priorities include:

• improving workflow robustness

• expanding automated tests

• improving documentation

• adding reproducible examples

• strengthening calibration validation

• refining the graphical interface

• improving packaging and release workflows

Installation

RosettaX requires Python 3.11 or later. CI checks Python 3.11, 3.12, and 3.13; the pinned PyMieSim release provides wheels for these versions.

Usage

After installation, launch RosettaX with:

rosettax

This starts the local graphical calibration application.

Common CLI options:

rosettax --host 0.0.0.0 --port 8050
rosettax --no-browser
rosettax --fcs-file-path /path/to/sample.fcs
rosettax --version

The application opens a browser based interface served from the local machine. It is intended for local calibration workflows and does not require a remote server.

Documentation

The documentation is available here:

RosettaX documentation

The documentation is expected to evolve with the application. At this stage, the README provides the high level overview, while the full documentation contains more detailed usage information.

Development

Install the project in editable mode with development dependencies:

pip install -e ".[dev,testing,documentation]"

Build a local PyInstaller bundle with:

pip install -e ".[bundling]"
pyinstaller rosettax.spec --clean

This creates a standalone bundle under dist/rosettax for the current operating system. Cross platform bundles must still be built natively on each target operating system.

Run the unit test suite with:

pytest

Run tests with coverage using:

pytest --cov=RosettaX

Run the integration suite in a separate process with real PyMieSim and Dash Bootstrap dependencies:

python -m pytest integration_tests -o addopts='' -v

The unit suite substitutes scientific and Bootstrap dependencies with stubs. The integration suite lives outside tests/ so it does not load those stubs; do not collect both suites in one pytest invocation. It checks native sphere and core-shell calculations, power scaling, Mie relation inversion, every registered page layout, and Dash HTTP endpoints. Missing dependencies or stub contamination fail the run instead of skipping checks. CI runs these checks on Python 3.11 and 3.13 on Linux, and Python 3.13 on macOS and Windows.

The repository includes configuration for testing, coverage, documentation, and continuous integration. These tools are intended to support reproducible development as the project matures.

Contributing

Contributions are welcome.

Useful contributions include:

• bug reports

• workflow suggestions

• documentation improvements

• test cases

• calibration examples

• user interface improvements

• support for additional calibration workflows

For substantial changes, please open an issue first so that the intended direction can be discussed before implementation.

Citation

If you use RosettaX in scientific work, please cite its Zenodo concept DOI: 10.5281/zenodo.19846629. It always resolves to the latest published Zenodo release.

@software{rosettax,
  author = {Poinsinet de Sivry-Houle, Martin},
  title = {RosettaX},
  year = {2026},
  publisher = {Zenodo},
  doi = {10.5281/zenodo.19846629},
  url = {https://doi.org/10.5281/zenodo.19846629}
}

The current citation and all archived versions are available at https://zenodo.org/records/19846629.

Contact

RosettaX is developed by Martin Poinsinet de Sivry Houle.

For collaboration, questions, or feedback:

martin.poinsinet.de.sivry@gmail.com

License

RosettaX is distributed under the GNU Affero General Public License v3.0 or later.

See the repository LICENSE file for the full license text.

Commercial licensing, support, validation, or custom deployment agreements may be discussed with the author.