Point spread function in interferometric scattering microscopy (iSCAT). Part I: aberrations in defocusing and axial...

Area of Science:

• Optical microscopy
• Nanotechnology
• Biophysics

Background:

• Interferometric scattering (iSCAT) microscopy is a label-free technique for detecting nanomatter.
• Previous iSCAT studies used Gaussian approximations for the point spread function (PSF).
• Accurate PSF modeling is crucial for nanoparticle tracking in complex environments and over extended axial ranges.

Purpose of the Study:

• To develop a quantitative vectorial diffraction model for the interferometric PSF (iPSF).
• To investigate the iPSF under various imaging conditions.
• To enable nanometric 3D localization of nanoparticles over extended axial ranges.

Main Methods:

• Vectorial diffraction modeling of the iPSF.
• Experimental iSCAT measurements.
• Finite-difference time-domain (FDTD) simulations.
• Analytical model fitting and unsupervised machine learning for localization.

Main Results:

• A robust vectorial diffraction model for the iPSF was established.
• The lateral shape of the iPSF encodes nanoparticle information.
• Nanometric 3D localization was achieved over a 10 µm axial range.
• Calibration-free machine learning approach demonstrated for localization.

Conclusions:

• The developed iPSF model accurately describes interferometric scattering microscopy.
• The findings enable precise 3D single particle tracking in complex scattering media.
• This advancement has significant implications for nanotechnology and biophysics research.