Two-photon fluorescence correlation spectroscopy with high count rates and low background using dielectric microspheres.
Two-photon excitation fluorescence is a powerful technique commonly used for biological imaging. However, the low absorption cross section of this non-linear process is a critical issue for performing biomolecular spectroscopy at the single molecule level. Enhancing the two-photon fluorescence signal would greatly improve the effectiveness of this technique, yet current methods struggle with medium enhancement factors and/or high background noise. Here, we show that the two-photon fluorescence signal from single Alexa Fluor 488 molecules can be enhanced up to 10 times by using a 3 µm diameter latex sphere while adding almost no photoluminescence background. We report a full characterization of the two-photon fluorescence enhancement by a single microsphere using fluorescence correlation spectroscopy. This opens new routes to enhance non-linear optical signals and extend biophotonic applications.
Heykel
H
Aouani
Peter
P
Schön
Sophie
S
Brasselet
Hervé
H
Rigneault
Jérôme
J
Wenger
Anal Chem. 2008 Sep 1;80(17):6800-4
18681458
Biophys J. 1995 Feb;68(2):694-701
7696520
Opt Express. 2004 Apr 5;12(7):1214-20
19474940
Appl Opt. 2001 Aug 20;40(24):3969-83
18360431
Nat Methods. 2005 Dec;2(12):932-40
16299478
Biophys J. 1999 Oct;77(4):2251-65
10512844
Science. 2003 Jan 31;299(5607):682-6
12560545
Opt Express. 2007 Apr 16;15(8):4935-42
19532742
Anal Chem. 2009 Jan 15;81(2):834-9
19099408
Opt Express. 2008 May 12;16(10):6930-40
18545397
Opt Express. 2009 Oct 12;17(21):19085-92
20372645
Opt Lett. 1995 Dec 15;20(24):2532
19865276
Opt Express. 2008 Mar 3;16(5):3008-20
18542387
Int J Mol Sci. 2010 Jan 13;11(1):206-21
20162011
Annu Rev Biomed Eng. 2000;2:399-429
11701518
Opt Express. 2005 Jan 24;13(2):526-33
19488381
Biophys J. 2005 Jun;88(6):4319-36
15792970
Opt Express. 2010 Jun 7;18(12):12807-17
20588409
Science. 1990 Apr 6;248(4951):73-6
2321027
Phys Rev Lett. 2005 Jan 14;94(1):017402
15698131
Opt Express. 2008 Sep 15;16(19):15297-303
18795067
Opt Express. 2008 Aug 18;16(17):12640-9
18711501
Opt Express. 2009 Feb 16;17(4):2089-94
19219113
Proc Natl Acad Sci U S A. 2000 Sep 12;97(19):10377-82
10973482
Proc Natl Acad Sci U S A. 1996 Oct 1;93(20):10763-8
8855254