A hybrid high-speed atomic force-optical microscope for visualizing single membrane proteins on eukaryotic cells

Nat Commun. 2013:4:2155. doi: 10.1038/ncomms3155.

Abstract

High-speed atomic force microscopy is a powerful tool for studying structure and dynamics of proteins. So far, however, high-speed atomic force microscopy was restricted to well-controlled molecular systems of purified proteins. Here we integrate an optical microscopy path into high-speed atomic force microscopy, allowing bright field and fluorescence microscopy, without loss of high-speed atomic force microscopy performance. This hybrid high-speed atomic force microscopy/optical microscopy setup allows positioning of the high-speed atomic force microscopy tip with high spatial precision on an optically identified zone of interest on cells. We present movies at 960 ms per frame displaying aquaporin-0 array and single molecule dynamics in the plasma membrane of intact eye lens cells. This hybrid setup allows high-speed atomic force microscopy imaging on cells about 1,000 times faster than conventional atomic force microscopy/optical microscopy setups, and allows first time visualization of unlabelled membrane proteins on a eukaryotic cell under physiological conditions. This development advances high-speed atomic force microscopy from molecular to cell biology to analyse cellular processes at the membrane such as signalling, infection, transport and diffusion.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Aquaporins / chemistry*
  • Aquaporins / metabolism
  • Biological Transport
  • Cell Membrane / metabolism
  • Cell Membrane / ultrastructure*
  • Cells, Immobilized
  • Escherichia coli / chemistry
  • Escherichia coli / ultrastructure*
  • Eye Proteins / chemistry*
  • Eye Proteins / metabolism
  • Lens, Crystalline / metabolism
  • Lens, Crystalline / ultrastructure*
  • Microscopy, Atomic Force / instrumentation
  • Microscopy, Atomic Force / methods*
  • Microscopy, Fluorescence / instrumentation
  • Microscopy, Fluorescence / methods*
  • Molecular Dynamics Simulation
  • Primary Cell Culture
  • Sheep
  • Sheep, Domestic
  • Time-Lapse Imaging

Substances

  • Aquaporins
  • Eye Proteins
  • aquaporin 0