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Applied Optics

Applied Optics


  • Vol. 41, Iss. 12 — Apr. 20, 2002
  • pp: 2285–2291

Simultaneous measurement of one- and two-photon excited fluorescence from a single sample: a detection method for oligonucleotides

Troy Alexander and Chieu D. Tran  »View Author Affiliations

Applied Optics, Vol. 41, Issue 12, pp. 2285-2291 (2002)

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A new method has been developed that is based on the use of a single-excitation wavelength from a cw laser to excite simultaneously one-photon and two-photon fluorescence (TPF). Fluorescence bands of a sample containing two oligonucleotides, one labeled with a one-photon fluorescence dye and the other with a TPF dye, can be measured simultaneously. The two fluorescence bands are well separated, because the one-photon excited fluorescence band is redshifted, whereas the TPF band is blueshifted from the excitation wavelength. The spectral separation was found to be as large as 200 nm when ADS 840NCS was used to label one oligonucleotide for one-photon fluorescence and Rhodamine Red-X dye was used for TPF. Spectral overlapping problem that plagues current DNA sequencing techniques can be eliminated effectively with this method.

© 2002 Optical Society of America

OCIS Codes
(000.1570) General : Chemistry
(120.0120) Instrumentation, measurement, and metrology : Instrumentation, measurement, and metrology
(140.0140) Lasers and laser optics : Lasers and laser optics
(170.6280) Medical optics and biotechnology : Spectroscopy, fluorescence and luminescence
(230.1040) Optical devices : Acousto-optical devices
(300.0300) Spectroscopy : Spectroscopy

Original Manuscript: July 27, 2001
Revised Manuscript: November 5, 2001
Published: April 20, 2002

Troy Alexander and Chieu D. Tran, "Simultaneous measurement of one- and two-photon excited fluorescence from a single sample: a detection method for oligonucleotides," Appl. Opt. 41, 2285-2291 (2002)

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