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Related Experiment Video

Updated: Feb 15, 2026

A Protocol for Real-time 3D Single Particle Tracking
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A Protocol for Real-time 3D Single Particle Tracking.

Shangguo Hou1, Kevin Welsher2

  • 1Department of Chemistry, Duke University.

Journal of Visualized Experiments : Jove
|January 25, 2018
PubMed
Summary
This summary is machine-generated.

This study introduces 3D Dynamic Photon Localization Tracking (3D-DyPLoT), a novel real-time three-dimensional single particle tracking system. It enables high-speed 3D particle tracking even at low photon rates, overcoming limitations of existing methods.

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Area of Science:

  • Biophysics
  • Optical Microscopy
  • Cellular Dynamics

Background:

  • Real-time three-dimensional single particle tracking (RT-3D-SPT) is crucial for studying fast cellular processes.
  • Existing RT-3D-SPT methods struggle with high-speed 3D particle tracking at low photon counts and are often complex to implement.

Purpose of the Study:

  • To present a robust and user-friendly RT-3D-SPT system named 3D Dynamic Photon Localization Tracking (3D-DyPLoT).
  • To enable high-speed (up to 20 µm²/s) and high-precision tracking of 3D diffusing particles at low photon count rates (down to 10 kHz).

Main Methods:

  • Utilizes a 2D electro-optic deflector (2D-EOD) and a tunable acoustic gradient (TAG) lens for dynamic 3D laser scanning.
  • Employs an optimized position estimation algorithm for accurate particle localization.
  • Features a simplified single excitation and detection path for ease of setup and robustness.

Main Results:

  • Demonstrates successful tracking of 110 nm fluorescent beads in water with high speed and precision.
  • Achieves tracking of particles with diffusive speeds up to 20 µm²/s at photon count rates as low as 10 kHz.
  • The 3D-DyPLoT system is shown to be robust and straightforward to assemble.

Conclusions:

  • 3D-DyPLoT offers a significant advancement for real-time 3D particle tracking in biological research.
  • The system's ease of implementation and performance at low photon rates make it widely applicable.
  • This protocol provides a detailed guide for building and utilizing the 3D-DyPLoT system.