Optimal 3D single-molecule localization for superresolution microscopy with aberrations and engineered point spread...

Area of Science:

• Biophysics
• Optical Microscopy
• Cell Biology

Background:

• Photo-activation localization microscopy (PALM/STORM) enables super-resolution imaging by precisely localizing sparse single molecules.
• Extension to 3D imaging offers new cellular exploration possibilities.
• Engineering the 3D point spread function (PSF) can theoretically enhance 3D localization precision.

Purpose of the Study:

• To present a general framework for an optimal 3D single-molecule localization estimator.
• To develop an estimator for noisy, aberrated, or engineered PSF imaging.
• To achieve efficient 3D localization precision, reaching the Cramer-Rao lower bound.

Main Methods:

• Implementation of a phase-retrieval enabled maximum-likelihood estimator.
• General framework applicable to various 3D PSF engineering strategies.
• Testing with experimental data from an engineered PSF microscope.

Main Results:

• The developed estimator is efficient, achieving the fundamental Cramer-Rao lower bound for x, y, and z localization precision.
• Demonstrated superior performance in low-photon-count 3D wide-field single-molecule localization.
• Validated the estimator's effectiveness with experimental data.

Conclusions:

• The phase-retrieval enabled maximum-likelihood estimator provides an optimal approach for 3D super-resolution microscopy.
• This method significantly improves 3D localization precision, especially under challenging imaging conditions.
• Unlocks potential for advanced cellular structure and function studies in 3D.