Real-Time 3D Single Particle Tracking: Towards Active Feedback Single Molecule Spectroscopy in Live Cells

Molecules : a Journal of Synthetic Chemistry and Natural Product Chemistry
Shangguo HouKevin Welsher

Abstract

Single molecule fluorescence spectroscopy has been largely implemented using methods which require tethering of molecules to a substrate in order to make high temporal resolution measurements. However, the act of tethering a molecule requires that the molecule be removed from its environment. This is especially perturbative when measuring biomolecules such as enzymes, which may rely on the non-equilibrium and crowded cellular environment for normal function. A method which may be able to un-tether single molecule fluorescence spectroscopy is real-time 3D single particle tracking (RT-3D-SPT). RT-3D-SPT uses active feedback to effectively lock-on to freely diffusing particles so they can be measured continuously with up to photon-limited temporal resolution over large axial ranges. This review gives an overview of the various active feedback 3D single particle tracking methods, highlighting specialized detection and excitation schemes which enable high-speed real-time tracking. Furthermore, the combination of these active feedback methods with simultaneous live-cell imaging is discussed. Finally, the successes in real-time 3D single molecule tracking (RT-3D-SMT) thus far and the roadmap going forward for this promising family of ...Continue Reading

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Citations

Jul 19, 2020·Nature Communications·Shangguo HouKevin Welsher
Oct 15, 2020·Advanced Healthcare Materials·Andrey S KlymchenkoNicolas Anton
Mar 13, 2021·Biophysical Journal·Yale E Goldman
Apr 17, 2021·The Journal of Chemical Physics·Maayan LevinYael Roichman
Jun 8, 2021·ACS Photonics·Olha MelnykRobert E Camley
Jan 18, 2020·Chemical Reviews·Shu-Lin LiuDai-Wen Pang

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Methods Mentioned

BETA
super-resolution microscopies
protein folding
FRET
fluorescence microscopy
fluorescence correlation spectroscopy

Software Mentioned

TSUNAMI
3D
DyPLoT
SPT
RT

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