Scattering workflow#

Use the scattering workflow when the calibration depends on an optical model rather than only on assigned reference intensities.

What is modeled#

RosettaX models scattering standards with PyMieSim using a Gaussian source and a photodiode detector configuration. The core optical inputs are:

  • wavelength

  • medium refractive index

  • particle refractive index, or core/shell refractive indices

  • detector numerical aperture

  • detector cache numerical aperture

  • blocker-bar numerical aperture

  • detector phi angle

  • detector gamma angle

  • detector sampling

Detector presets can also contribute angular weights and effective geometry corrections before the coupling values are computed.

Supported cytometer presets#

RosettaX currently ships detector presets for these grouped instrument families:

  • BD Biosciences: FACSCanto II - FSC, FACSCanto II - SSC

  • Beckman Coulter Life Sciences: CytoFLEX - Fluorescence, CytoFLEX - SSC

  • Custom: Generic detector

  • Apogee: Forward, Side

In the scattering workflow, the optical preset selector is split into brand and model. Choose a brand first, then choose the supported model under that brand. The selected model still resolves to the same saved detector preset name inside the calibration payload.

If your company would like RosettaX to support another cytometer, contact the maintainer with an example FCS file, the detector naming you want recognized, and any public optical-geometry documentation that should inform the preset.

For the full per-system detector geometry overview (one card per cytometer system with 3D figures and placeholders), see Flow-cytometer systems.

How the calibration-standard relation is built#

RosettaX parses the calibration-standard table, keeps only rows with finite diameter and measured-peak values, computes modeled coupling for those standard particles, and then fits the measured peaks against those theoretical coupling values.

The current instrument-response model is linear:

\[\mathrm{theoretical\_coupling} = m \cdot \mathrm{measured\_peak} + b\]

The default behavior is to force the intercept to zero. RosettaX stores the fit parameters and R^2 in the instrument_response block.

Material refractive indices#

Material presets such as water, PBS, polystyrene, silica, PMMA, and lipid-like media are resolved through the packaged Sellmeier bank at the selected wavelength. Numeric presets remain numeric. This means the refractive indices used for the fit can change with wavelength and are not treated as hard-coded constants.

Saved payload#

The scattering payload preserves both the fit and the modeled standard context:

  • source channel and output quantity

  • instrument-response fit parameters

  • the calibration-standard Mie relation

  • the reference table used for fitting

  • metadata needed to reconstruct the optical assumptions later

That payload is what allows RosettaX to apply a scatter calibration later by first converting measured peak to estimated coupling and then converting that coupling onto a target Mie relation.