Fluorescence imaging with one-nanometer accuracy (fiona)

Yong Wang, E. Cai, Janet Sheung, Sang Hak Lee, Kai Wen Teng, Paul R. Selvin

Research output: Contribution to journalArticle

Abstract

Fluorescence imaging with one-nanometer accuracy (FIONA) is a simple but useful technique for localizing single fluorophores with nanometer precision in the x-y plane. Here a summary of the FIONA technique is reported and examples of research that have been performed using FIONA are briefly described. First, how to set up the required equipment for FIONA experiments, i.e., a total internal reflection fluorescence microscopy (TIRFM), with details on aligning the optics, is described. Then how to carry out a simple FIONA experiment on localizing immobilized Cy3-DNA single molecules using appropriate protocols, followed by the use of FIONA to measure the 36 nm step size of a single truncated myosin Va motor labeled with a quantum dot, is illustrated. Lastly, recent effort to extend the application of FIONA to thick samples is reported. It is shown that, using a water immersion objective and quantum dots soaked deep in sol-gels and rabbit eye corneas (>200 µm), localization precision of 2-3 nm can be achieved.

Original languageEnglish (US)
Article numbere51774
JournalJournal of Visualized Experiments
Issue number91
DOIs
StatePublished - Sep 26 2014

Keywords

  • FIONA
  • Fluorescence imaging
  • Issue 91
  • Molecular Biology
  • Myosin walking
  • Nanometer precision
  • Thick tissue

ASJC Scopus subject areas

  • Neuroscience(all)
  • Chemical Engineering(all)
  • Biochemistry, Genetics and Molecular Biology(all)
  • Immunology and Microbiology(all)

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