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Mass-Sensitive Particle Tracking to Characterize Membrane-Associated Macromolecule Dynamics

Mass-Sensitive Particle Tracking to Characterize Membrane-Associated Macromolecule Dynamics

Published on: February 18, 2022

Mass-Sensitive Particle Tracking to Characterize Membrane-Associated Macromolecule Dynamics.

Frederik Steiert¹, Tamara Heermann², Nikolas Hundt³

¹Department of Cellular and Molecular Biophysics, Max Planck Institute of Biochemistry; Department of Physics, Technical University Munich.

Journal of Visualized Experiments : Jove

|March 7, 2022

Summary

Mass-sensitive particle tracking (MSPT) quantifies transient macromolecule interactions with lipid membranes in real-time. This technique, using interferometric scattering microscopy-based mass photometry (iSCAT-MP), reveals diffusion and mass of membrane-associated proteins.

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Area of Science:

Biophysics
Cell Biology
Biochemistry

Background:

Transient macromolecule interactions with lipid membranes are crucial for biological processes but difficult to study.
Standard biophysical methods struggle to analyze these dynamic membrane events.
Interferometric scattering microscopy (iSCAT)-based mass photometry (MP) offers a novel solution.

Purpose of the Study:

To present a detailed protocol for mass-sensitive particle tracking (MSPT) of membrane-associated systems.
To enable real-time quantitative analysis of macromolecule diffusion and mass on biological membranes.
To demonstrate the utility of MSPT for studying membrane-catalyzed dynamics.

Main Methods:

Utilizing interferometric scattering microscopy (iSCAT)-based mass photometry (MP).
Optimizing background removal for two-dimensional particle motion analysis.
Applying mass-sensitive particle tracking (MSPT) to unlabeled macromolecules on membranes.

Main Results:

Achieved millisecond time resolution for membrane-associated particle tracking.
Demonstrated a mass detection limit down to 50 kDa, depending on the MP system.
Successfully analyzed the dynamics of membrane interactors like annexin V.

Conclusions:

MSPT provides a powerful tool for quantitative characterization of transient membrane interactions.
This technique allows real-time analysis of diffusion and molecular mass of membrane-bound macromolecules.
MSPT significantly advances the study of membrane-associated macromolecule dynamics.