Subdiffraction-limit two-photon fluorescence microscopy for GFP-tagged cell imaging

Biophys J. 2009 Dec 16;97(12):3224-8. doi: 10.1016/j.bpj.2009.09.038.

Abstract

We report applications of two-photon excitation fluorescence (2PEF) microscopy with subdiffraction-limit resolution for green-fluorescent-protein-tagged cell imaging. The microscope integrates 2PEF microscopy and stimulated emission depletion microscopy in one microscope that has the benefits of both techniques: intrinsic three-dimensional resolution, confined photobleaching, and subdiffraction-limit resolution. The subdiffraction-limit resolution was demonstrated by resolving green-fluorescent-protein-tagged caveolar vesicles located within a distance shorter than the diffraction limit of a regular 2PEF microscope, which is approximately 250 nm even with the best optics. The full width at half-maximum of the effective point-spread function for the 2PEF microscope was estimated to be approximately 54 nm.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • CHO Cells
  • Cricetinae
  • Cricetulus
  • Green Fluorescent Proteins / metabolism*
  • Microscopy, Fluorescence / methods*
  • Molecular Imaging / methods*
  • Photons*

Substances

  • Green Fluorescent Proteins