Fluorescence Imaging with One-Nanometer Accuracy (FIONA) of Cy3-DNA under Deoxygenation Conditions
- Paul R. Selvin1,
- Tyler Lougheed,
- Melinda Tonks Hoffman,
- Hyokeun Park,
- Hamza Balci,
- Benjamin H. Blehm and
- Erdal Toprak
- ↵1Corresponding author (selvin{at}uiuc.edu)
INTRODUCTION
FIONA, short for fluorescence imaging with one-nanometer accuracy, is a simple method for achieving localization of single (or single groups of) fluorophores with nanometer accuracy in the xy plane. This protocol provides the steps necessary to run a control experiment, using DNA labeled with Cy3, to assess the efficacy of the FIONA setup and the level of attainable resolution. The Cy3-DNA is immobilized on a coverslip and imaged under deoxygenation conditions. It is important that the fluorophores remain photostable throughout the experiment. This requires an oxygen-scavenging system (e.g., glucose oxidase and catalase) in the medium.
