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Optimizing the metric in sensorless adaptive optical microscopy with fluorescence fluctuations.

Joseph Gallagher, Antoine Delon, Philippe Moreau

|August 10, 2017

Summary

Sensorless adaptive optics (AO) correction for fluorescence microscopy requires optimal metric selection. Molecular brightness is better for few, bright fluorophores; count rate is superior for concentrated solutions and large aberrations.

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Area of Science:

Biophysics
Optical Engineering
Microscopy

Background:

Adaptive optics (AO) corrects optical aberrations in microscopy.
Sensorless AO methods optimize corrections without external wavefront sensors.
Fluorescence fluctuations microscopy (FFM) quantifies molecular dynamics but is sensitive to aberrations.

Purpose of the Study:

To evaluate fluorescence count rate and molecular brightness as metrics for sensorless AO in FFM.
To compare their noise, sensitivity to aberrations, and correction accuracy.
To develop a theoretical framework for predicting AO correction accuracy.

Main Methods:

Experimental measurements in 3D fluorophore solutions.
Analysis of metric noise and aberration sensitivity.
Assessment of AO correction accuracy using residual aberrations.
Theoretical modeling of correction accuracy.

Main Results:

Molecular brightness metric yields more accurate corrections for sparse, bright fluorophores.
Count rate metric performs better in concentrated solutions and for large aberrations.
A theoretical framework accurately predicts correction accuracy based on metric properties.

Conclusions:

Metric choice critically impacts sensorless AO performance in FFM.
Count rate is a more robust metric for correcting significant aberrations.
Understanding metric characteristics enables optimized AO strategies for FFM.