Single-Molecule Detection of a Fluorescent Nucleobase Analogue via Multiphoton Excitation

David Nobis, Rachel S. Fisher, Mats Simmermacher, Patrycja A. Hopkins, Yitzhak Tor, Anita C. Jones, Steven W. Magennis

Research output: Contribution to journalArticlepeer-review

Abstract

The ability to routinely detect fluorescent nucleobase analogues at the single-molecule level would create a wealth of opportunities to study nucleic acids. We report the multiphoton-induced fluorescence and single-molecule detection of a dimethylamine-substituted extended-6-aza-uridine (DMAthaU). We show that DMAthaU can exist in a highly fluorescent form, emitting strongly in the visible region (470–560 nm). Using pulse-shaped broadband Ti:sapphire laser excitation, DMAthaU undergoes two-photon (2P) absorption at low excitation powers, switching to three-photon (3P) absorption at high incident intensity. The assignment of a 3P process is supported by cubic response calculations. Under both 2P and 3P excitation, the single-molecule brightness was over an order of magnitude higher than reported previously for any fluorescent base analogue, which facilitated the first single-molecule detection of an emissive nucleoside with multiphoton excitation.
Original languageEnglish
Pages (from-to)5008-5012
JournalThe Journal of Physical Chemistry Letters
Early online date9 Aug 2019
DOIs
Publication statusE-pub ahead of print - 9 Aug 2019

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