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Peptide-PAINT Using a Transfected-Docker Enables Live- and Fixed-Cell Super-Resolution Imaging.

Barun Kumar Maity1, Duncan Nall1, Yongjae Lee2

  • 1Department of Physics, University of Illinois at Urbana-Champaign, Urbana, 61801, USA.

Small Methods
|February 3, 2023
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Summary

This study introduces a new method for super-resolution microscopy called Peptide-PAINT. It enables high-resolution imaging of live cells and reveals nanoscale details of cellular structures.

Keywords:
Golgi bodylive cellspeptide-PAINTsingle-molecule imagingsuper-resolutiontransfected-docker

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Area of Science:

  • Biophysics
  • Cell Biology
  • Microscopy

Background:

  • Point accumulation for imaging in nanoscale topography (PAINT) is a super-resolution microscopy technique achieving 5-25 nm resolution using DNA or peptide tethers.
  • Current methods typically involve external addition of dockers, limiting applications in live cells.

Purpose of the Study:

  • To develop an improved PAINT technique for super-resolution microscopy.
  • To enable live-cell imaging of surface proteins in mammalian cells and neurons.
  • To achieve higher resolution and simplify experimental procedures.

Main Methods:

  • Transfecting proteins of interest with a docker-coil for PAINT imaging.
  • Applying Peptide-PAINT to live mammalian cells and neurons.
  • Imaging organelles in fixed cells to analyze ultrastructural details.

Main Results:

  • Achieved a localization precision of approximately 10 nm, comparable to standard PAINT.
  • Enabled Peptide-PAINT application in live cells under physiological conditions.
  • Resolved nanoscale features, including 40-60 nm repeats in vimentin filaments and sub-100 nm histone-rich regions.

Conclusions:

  • Transfected dockers offer experimental advantages and simplifications for PAINT super-resolution microscopy.
  • Peptide-PAINT is a viable technique for high-resolution live-cell imaging of surface proteins.
  • The method provides unprecedented insights into cellular ultrastructure.