Theoretical comparison of real-time single-particle tracking techniques

Bertus van Heerden; Tjaart P. J. Krüger

doi:10.1117/12.2582974

5 March 2021 Theoretical comparison of real-time single-particle tracking techniques

Bertus van Heerden, Tjaart P. J. Krüger

Proceedings Volume 11650, Single Molecule Spectroscopy and Superresolution Imaging XIV; 116500K (2021) https://doi.org/10.1117/12.2582974
Event: SPIE BiOS, 2021, Online Only

Abstract

One of the main challenges in studying single biomolecules in a native or near-native environment is their constant diffusive motion. A method capable of keeping a single biomolecule in the detection volume without disturbing its surrounding environment is real-time single-particle tracking (SPT). There are a number of real-time SPT methods in use, and comparisons in literature are usually made based on experimental results, which, due to variations in experimental setups and specimens, are of limited use. In this study, we used statistical calculations to compare the theoretical performance of different localization methods, namely the orbital method, the Knight’s Tour (grid scan) method and MINFLUX, in the context of both fluorescence-based and interferometric scattering (iSCAT) approaches. While the Knight’s Tour method is able to track the fastest diffusion, MINFLUX is able to achieve the greatest precision, albeit only for slowly diffusing particles. For comparing iSCAT and fluorescence, we considered different biological examples, including both intrinsically fluorescent particles and particles with a fluorescent label. The relative success of iSCAT vs fluorescence is strongly dependent on the particle size, photophysical properties of the fluorophore, and the fluorophore density. This work should serve as a guideline for choosing a suitable method for experimental implementation of real-time SPT based on the sample of interest. The analysis used can easily be extended to other methods of interest that we have not considered.

Conference Presentation

Citation Download Citation

Bertus van Heerden and Tjaart P. J. Krüger "Theoretical comparison of real-time single-particle tracking techniques", Proc. SPIE 11650, Single Molecule Spectroscopy and Superresolution Imaging XIV, 116500K (5 March 2021); https://doi.org/10.1117/12.2582974

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