K Estimator Validation — Fixed Illumination, Variable Bead Size

This example demonstrates how to estimate the K parameter, which characterizes how the absolute signal noise (Robust STD) scales with the square root of the median signal when varying bead size under fixed illumination power.

Setup and configuration

import numpy as np
from TypedUnit import ureg

from FlowCyPy import FlowCytometer, SimulationSettings
from FlowCyPy.calibration import KEstimator
from FlowCyPy.fluidics import FlowCell, Fluidics, ScattererCollection
from FlowCyPy.opto_electronics import (
    Detector,
    OptoElectronics,
    TransimpedanceAmplifier,
    source,
)
from FlowCyPy.signal_processing import Digitizer, SignalProcessing

Configure simulation-level noise assumptions

SimulationSettings.include_noises = True
SimulationSettings.include_shot_noise = True
SimulationSettings.include_dark_current_noise = False
SimulationSettings.include_source_noise = False
SimulationSettings.include_amplifier_noise = False
SimulationSettings.assume_perfect_hydrodynamic_focusing = True
SimulationSettings.assume_amplifier_bandwidth_is_infinite = True
SimulationSettings.assume_perfect_digitizer = True
SimulationSettings.evenly_spaced_events = True

np.random.seed(3)  # Reproducibility

Construct simulation components

flow_cell = FlowCell(
    sample_volume_flow=80 * ureg.microliter / ureg.minute,
    sheath_volume_flow=1 * ureg.milliliter / ureg.minute,
    width=400 * ureg.micrometer,
    height=400 * ureg.micrometer,
)

scatterer_collection = ScattererCollection(medium_refractive_index=1.33 * ureg.RIU)

fluidics = Fluidics(scatterer_collection=scatterer_collection, flow_cell=flow_cell)

_source = source.GaussianBeam(
    numerical_aperture=0.2 * ureg.AU,
    wavelength=450 * ureg.nanometer,
    optical_power=150 * ureg.milliwatt,  # Fixed illumination power
)

digitizer = Digitizer(
    bit_depth="16bit",
    saturation_levels=(0 * ureg.volt, 2 * ureg.volt),
    sampling_rate=60 * ureg.megahertz,
)

amplifier = TransimpedanceAmplifier(
    gain=10 * ureg.volt / ureg.ampere,
    bandwidth=60 * ureg.megahertz,
)

detector_0 = Detector(
    name="default",
    phi_angle=0 * ureg.degree,
    numerical_aperture=0.2 * ureg.AU,
    cache_numerical_aperture=0.0 * ureg.AU,
    responsivity=1 * ureg.ampere / ureg.watt,
)

opto_electronics = OptoElectronics(
    detectors=[detector_0], source=_source, amplifier=amplifier
)

signal_processing = SignalProcessing(
    digitizer=digitizer,
    analog_processing=[],
)

flow_cytometer = FlowCytometer(
    opto_electronics=opto_electronics,
    fluidics=fluidics,
    signal_processing=signal_processing,
    background_power=_source.optical_power * 0.001,
)

Run K Estimation Simulation

k_estimator = KEstimator(debug_mode=False)

k_estimator.add_batch(
    bead_diameters=np.linspace(300, 900, 15) * ureg.nanometer,
    illumination_power=_source.optical_power,
    flow_cytometer=flow_cytometer,
    particle_count=50 * ureg.particle,
)

[INFO] Simulating bead 1/15: 300.0 nanometer
[INFO] Simulating bead 2/15: 342.85714285714283 nanometer
[INFO] Simulating bead 3/15: 385.7142857142857 nanometer
[INFO] Simulating bead 4/15: 428.57142857142856 nanometer
[INFO] Simulating bead 5/15: 471.42857142857144 nanometer
[INFO] Simulating bead 6/15: 514.2857142857142 nanometer
[INFO] Simulating bead 7/15: 557.1428571428571 nanometer
[INFO] Simulating bead 8/15: 600.0 nanometer
[INFO] Simulating bead 9/15: 642.8571428571429 nanometer
[INFO] Simulating bead 10/15: 685.7142857142857 nanometer
[INFO] Simulating bead 11/15: 728.5714285714286 nanometer
[INFO] Simulating bead 12/15: 771.4285714285713 nanometer
[INFO] Simulating bead 13/15: 814.2857142857142 nanometer
[INFO] Simulating bead 14/15: 857.1428571428571 nanometer
[INFO] Simulating bead 15/15: 900.0 nanometer

Plot estimation

k_estimator.plot()

<Figure size 800x500 with 1 Axes>

Plot relevant statistics

k_estimator.plot_statistics()

<Figure size 800x500 with 2 Axes>