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Related Experiment Video

Updated: Aug 2, 2025

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

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BNP-Track: A framework for superresolved tracking.

Ioannis Sgouralis1, Lance W Q Xu 徐伟青2,3, Ameya P Jalihal4

  • 1Department of Mathematics, University of Tennessee, Knoxville, TN 37996, USA.

Biorxiv : the Preprint Server for Biology
|April 17, 2023
PubMed
Summary

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This study introduces a new superresolution method for tracking multiple, closely spaced single molecules in dynamic biological samples. It achieves high accuracy without relying on photophysics, enabling real-time observation of molecular processes.

Area of Science:

  • Biophysics
  • Molecular Imaging
  • Cell Biology

Background:

  • Superresolution microscopy methods like STORM, PALM, and PAINT offer nanoscale localization but are limited to static samples due to reliance on rare photophysical events.
  • Existing methods struggle to distinguish and track multiple, closely spaced emitters simultaneously, especially in dynamic biological environments.

Approach:

  • Leverages spatiotemporal correlations in dynamic widefield imaging data to enable superresolution tracking of multiple emitters.
  • Develops a fully joint Bayesian nonparametric posterior distribution for simultaneous emitter number determination, localization, and linking across frames.
  • Avoids traditional tracking paradigms that separate emitter number determination, localization, and linking tasks.

Key Points:

  • Achieves superresolution tracking of simultaneously active, closely spaced emitters without photodynamic reliance.
Keywords:
Bayesian nonparametricsFluorescence microscopyImage analysisNanoscopySingle molecule trackingSuperresolution microscopy

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Last Updated: Aug 2, 2025

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  • Determines emitter numbers and tracks with localization accuracy comparable to widefield superresolution (≈50nm) per frame.
  • Quantifies uncertainty from various noise sources including shot noise, camera artifacts, and motion.

    • Conclusions:

    • Extends superresolution microscopy to simultaneously track multiple dynamic single molecules below the diffraction limit.
    • Demonstrates accuracy and robustness for both *in cellulo* and synthetic data, outperforming existing tools in crowded conditions.
    • Provides a powerful new tool for observing life at the molecular level in real-time.