Jump to main content
Jump to site search

Issue 45, 2018
Previous Article Next Article

Pulse-shaped two-photon excitation of a fluorescent base analogue approaches single-molecule sensitivity

Author affiliations

Abstract

Fluorescent nucleobase analogues (FBAs) have many desirable features in comparison to extrinsic fluorescent labels, but they are yet to find application in ultrasensitive detection. Many of the disadvantages of FBAs arise from their short excitation wavelengths (often in the ultraviolet), making two-photon excitation a potentially attractive approach. Pentacyclic adenine (pA) is a recently developed FBA that has an exceptionally high two-photon brightness. We have studied the two-photon-excited fluorescence properties of pA and how they are affected by incorporation in DNA. We find that pA is more photostable under two-photon excitation than via resonant absorption. When incorporated in an oligonucleotide, pA has a high two-photon cross section and emission quantum yield, varying with sequence context, resulting in the highest reported brightness for such a probe. The use of a two-photon microscope with ultrafast excitation and pulse shaping has allowed the detection of pA-containing oligonucleotides in solution with a limit of detection of ∼5 molecules, demonstrating that practical single-molecule detection of FBAs is now within reach.

Graphical abstract: Pulse-shaped two-photon excitation of a fluorescent base analogue approaches single-molecule sensitivity

Back to tab navigation

Supplementary files

Article information


Submitted
29 Aug 2018
Accepted
30 Oct 2018
First published
30 Oct 2018

This article is Open Access

Phys. Chem. Chem. Phys., 2018,20, 28487-28498
Article type
Paper

Pulse-shaped two-photon excitation of a fluorescent base analogue approaches single-molecule sensitivity

R. S. Fisher, D. Nobis, A. F. Füchtbauer, M. Bood, M. Grøtli, L. M. Wilhelmsson, A. C. Jones and S. W. Magennis, Phys. Chem. Chem. Phys., 2018, 20, 28487
DOI: 10.1039/C8CP05496G

This article is licensed under a Creative Commons Attribution 3.0 Unported Licence. Material from this article can be used in other publications provided that the correct acknowledgement is given with the reproduced material.

Reproduced material should be attributed as follows:

  • For reproduction of material from NJC:
    [Original citation] - Published by The Royal Society of Chemistry (RSC) on behalf of the Centre National de la Recherche Scientifique (CNRS) and the RSC.
  • For reproduction of material from PCCP:
    [Original citation] - Published by the PCCP Owner Societies.
  • For reproduction of material from PPS:
    [Original citation] - Published by The Royal Society of Chemistry (RSC) on behalf of the European Society for Photobiology, the European Photochemistry Association, and RSC.
  • For reproduction of material from all other RSC journals:
    [Original citation] - Published by The Royal Society of Chemistry.

Information about reproducing material from RSC articles with different licences is available on our Permission Requests page.


Social activity

Search articles by author

Spotlight

Advertisements