Small quantum dots conjugated to nanobodies as immunofluorescence probes for nanometric microscopy

Bioconjug Chem. 2014 Dec 17;25(12):2205-11. doi: 10.1021/bc5004179. Epub 2014 Nov 25.

Abstract

Immunofluorescence, a powerful technique to detect specific targets using fluorescently labeled antibodies, has been widely used in both scientific research and clinical diagnostics. The probes should be made with small antibodies and high brightness. We conjugated GFP binding protein (GBP) nanobodies, small single-chain antibodies from llamas, with new ∼7 nm quantum dots. These provide simple and versatile immunofluorescence nanoprobes with nanometer accuracy and resolution. Using the new probes we tracked the walking of individual kinesin motors and measured their 8 nm step sizes; we tracked Piezo1 channels, which are eukaryotic mechanosensitive channels; we also tracked AMPA receptors on living neurons. Finally, we used a new super-resolution algorithm based on blinking of (small) quantum dots that allowed ∼2 nm precision.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Algorithms
  • Cell Membrane / metabolism
  • Green Fluorescent Proteins / chemistry
  • Green Fluorescent Proteins / genetics
  • Ion Channels / analysis
  • Ion Channels / genetics
  • Ion Channels / metabolism
  • Kinesins / analysis
  • Kinesins / metabolism
  • Microscopy, Electron, Transmission
  • Microscopy, Fluorescence / methods*
  • Microtubules / metabolism
  • Molecular Probes / chemistry
  • Neurons / metabolism
  • Quantum Dots / chemistry*
  • Receptors, AMPA / analysis
  • Receptors, AMPA / metabolism
  • Single-Chain Antibodies / chemistry
  • Single-Domain Antibodies / chemistry*

Substances

  • Ion Channels
  • Molecular Probes
  • PIEZO1 protein, human
  • Receptors, AMPA
  • Single-Chain Antibodies
  • Single-Domain Antibodies
  • Green Fluorescent Proteins
  • Kinesins