Monomeric Garnet, a far-red fluorescent protein for live-cell STED imaging

Sci Rep. 2015 Dec 9;5:18006. doi: 10.1038/srep18006.

Abstract

The advancement of far-red emitting variants of the green fluorescent protein (GFP) is crucially important for imaging live cells, tissues and organisms. Despite notable efforts, far-red marker proteins still need further optimization to match the performance of their green counterparts. Here we present mGarnet, a robust monomeric marker protein with far-red fluorescence peaking at 670 nm. Thanks to its large extinction coefficient of 95,000 M(-1)cm(-1), mGarnet can be efficiently excited with 640-nm light on the red edge of its 598-nm excitation band. A large Stokes shift allows essentially the entire fluorescence emission to be collected even with 640-nm excitation, counterbalancing the lower fluorescence quantum yield of mGarnet, 9.1%, that is typical of far-red FPs. We demonstrate an excellent performance as a live-cell fusion marker in STED microscopy, using 640 nm excitation and 780 nm depletion wavelengths.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • COS Cells
  • Chlorocebus aethiops
  • Gene Expression*
  • Green Fluorescent Proteins
  • Luminescent Proteins / chemistry
  • Luminescent Proteins / genetics*
  • Luminescent Proteins / metabolism*
  • Microscopy, Fluorescence
  • Molecular Imaging / methods*
  • Protein Engineering
  • Recombinant Fusion Proteins

Substances

  • Luminescent Proteins
  • Recombinant Fusion Proteins
  • red fluorescent protein
  • Green Fluorescent Proteins