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Protein Dynamics in Living Cells01:19

Protein Dynamics in Living Cells

Different fluorescence-based techniques are used to study the protein dynamics in living cells. These techniques include FRAP, FRET, and PET.
Fluorescent recovery after photobleaching (FRAP) is a fluorescent-protein-based detection technique used to quantify protein movement rates within the cell. This method exposes a small portion of the cell to an intense laser beam. The laser beam causes permanent photobleaching of the fluorophore-tagged proteins in the exposed region. As the bleached...

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Mapping Molecular Diffusion in the Plasma Membrane by Multiple-Target Tracing (MTT)
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Mapping Molecular Diffusion in the Plasma Membrane by Multiple-Target Tracing (MTT)

Published on: May 27, 2012

Particle tracking single protein-functionalized quantum dot diffusion and binding at silica surfaces.

Jack C Rife1, James P Long, John Wilkinson

  • 1Naval Research Laboratory, Washington, D.C. 20375, USA. rife@nrl.navy.mil

Langmuir : the ACS Journal of Surfaces and Colloids
|August 27, 2009
PubMed
Summary

We studied quantum dots (QDs) for tracking at surfaces. While QDs diffused well on smooth silica, rougher surfaces caused transient binding, indicating adsorption to defects.

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Published on: October 23, 2014

Area of Science:

  • Biophysics
  • Materials Science
  • Surface Chemistry

Background:

  • Single-particle tracking microscopy is crucial for observing molecular dynamics.
  • Quantum dots (QDs) offer bright, photostable fluorescence for tracking applications.
  • Understanding QD behavior at surfaces is essential for reliable microscopy.

Purpose of the Study:

  • To evaluate commercial streptavidin-functionalized QDs for surface-based single-particle tracking.
  • To measure QD diffusion and nonspecific binding kinetics on silica surfaces under varying conditions.
  • To characterize QD properties relevant for optimizing tracking experiments.

Main Methods:

  • Total internal reflectance fluorescence (TIRF) microscopy was employed.
  • Single QD diffusion and adsorption were analyzed on silica surfaces.
  • QD parameters like hydrodynamic radius, charge, and optical properties were assessed.

Main Results:

  • QD diffusion coefficients approached bulk solution values on smooth, repulsive silica surfaces.
  • Transient adsorptions were observed on rougher silica surfaces, attributed to defect sites.
  • Binding energies were estimated from adsorption lifetimes.
  • QD optical properties, including blinking and bleaching, were characterized.

Conclusions:

  • Streptavidin-QDs exhibit predictable diffusion on ideal silica surfaces.
  • Surface imperfections and adsorbates significantly influence QD binding.
  • QD surface interactions must be considered for accurate single-particle tracking at interfaces.