1. Academic Validation
  2. Synthesis of Janelia Fluor HaloTag and SNAP-Tag Ligands and Their Use in Cellular Imaging Experiments

Synthesis of Janelia Fluor HaloTag and SNAP-Tag Ligands and Their Use in Cellular Imaging Experiments

  • Methods Mol Biol. 2017;1663:179-188. doi: 10.1007/978-1-4939-7265-4_15.
Jonathan B Grimm 1 Timothy A Brown 1 Brian P English 1 Timothée Lionnet 1 Luke D Lavis 2
Affiliations

Affiliations

  • 1 Janelia Research Campus, Howard Hughes Medical Institute, Ashburn, VA, USA.
  • 2 Janelia Research Campus, Howard Hughes Medical Institute, Ashburn, VA, USA. lavisl@janelia.hhmi.org.
Abstract

The development of genetically encoded self-labeling protein tags such as the HaloTag and SNAP-tag has expanded the utility of chemical dyes in microscopy. Intracellular labeling using these systems requires small, cell-permeable dyes with high brightness and photostability. We recently discovered a general method to improve the properties of classic fluorophores by replacing N,N-dimethylamino groups with four-membered azetidine rings to create the "Janelia Fluor" dyes. Here, we describe the synthesis of the HaloTag and SNAP-tag ligands of Janelia Fluor 549 and Janelia Fluor 646 as well as standard labeling protocols for use in ensemble and single-molecule cellular imaging.

Keywords

Fluorescence microscopy; Fluorophore; HaloTag; Organic chemistry; SNAP-tag; Single-molecule imaging.

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