Global fitting for high-accuracy multi-channel single-molecule localization

Abstract

Multi-channel detection in single-molecule localization microscopy greatly increases information content for various biological applications. Here, we present globLoc, a graphics processing unit based global fitting algorithm with flexible PSF modeling and parameter sharing, to extract maximum information from multi-channel single molecule data. As signals in multi-channel data are highly correlated, globLoc links parameters such as 3D coordinates or photon counts across channels, improving localization precision and robustness. We show, both in simulations and experiments, that global fitting can substantially improve the 3D localization precision for biplane and 4Pi single-molecule localization microscopy and color assignment for ratiometric multicolor imaging.

Field dependent deep learning enables high-throughput whole-cell 3D super-resolution imaging

“…1b ). Second, we integrated globLoc with flexible parameter sharing 26 functionality to our vectorial PSF fitter which could link parameters (e.g. aberrations, xyz positions, photons, background) among different single molecule images.…”

“…MLE fitting routine was employed to analyze the bead stacks with Zernike coefficients as the fitting parameters. We then employed globLoc 26 for the parameter estimation. Each image in the bead stack can be regarded as an individual channel, whose fitting parameters can be shared or fit independently.…”

“…Since vectorial PSF fitter is computational challenging, we implemented the fitting pipeline using CUDA C/C++ in NVIDIA CUDA® enabled graphic cards to enable fast modeling of the spatially variant PSFs in the large FOV. The framework of the MLE fitting method follows the previous work 26 which employed a modified Levenberg-Marquardt iterative schemes for non-linear optimization. To accurately retrieve the aberrations from point emitter, single molecule images at different z positions are normally needed.…”

“…Since vectorial PSF fitter is computational challenging, we implemented the fitting pipeline using CUDA C/C++ in NVIDIA CUDA® enabled graphic cards to enable fast modeling of the spatially variant PSFs in the large FOV. The framework of the MLE fitting method follows the previous work 26…”

Obtaining 3D super-resolution images by utilizing rotationally symmetric structures and 2D-to-3D transformation

“…There are several single-molecule approaches to gain Z-dimensional data by capturing and contorting the emission PSF [17] , [39] , [40] , [41] . One such technique captures the emission PSFs at multiple focal planes by multiple detectors, known multifocal or bi-plane imaging.…”