Abstract

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

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  1. L. M. Smith, J. Z. Sander, R. J. Kaiser, P. Hughes, C. Dodd, C. R. Connell, C. Heiner, S. B. H. Kent, L. E. Hood, “Fluorescence detection in automated DNA sequencing analysis,” Nature 321, 674–679 (1986).
    [CrossRef] [PubMed]
  2. L. M. Smith, “The future of DNA sequencing,” Science 262, 530–532 (1993).
    [CrossRef] [PubMed]
  3. S. M. Clark, R. A. Mathies, “Multiplex dsDNA fragment sizing using dimeric intercalation dyes and capillary array electrophoresis: ionic effects on the stability and electrophoretic mobility of DNA-dye complexes,” Anal. Chem. 69, 1355–1363 (1997).
    [CrossRef] [PubMed]
  4. S. A. Soper, B. L. Legendre, D. C. Williams, “On-line fluorescence lifetime determinations in capillary electrophoresis,” Anal. Chem. 67, 4358–4365 (1995).
    [CrossRef] [PubMed]
  5. H. He, L. B. McGown, “DNA sequencing by capillary electrophoresis with four-decay fluorescence detection,” Anal. Chem. 72, 5865–5873 (2000).
    [CrossRef]
  6. H. He, B. K. Nunnally, L. Li, L. B. McGown, “On-the-fly fluorescence lifetime detection of dye labeled DNA primers for multiplex analysis,” Anal. Chem. 70, 3413–3418 (1998).
    [CrossRef] [PubMed]
  7. L. G. Lee, S. L. Spergeon, C. R. Heiner, S. C. Benson, B. B. Resenblum, S. M. Menchen, R. J. Graham, A. Constantinescu, K. G. Upadhya, J. M. Cassel, “New energy transfer dyes for DNA sequencing,” Nucleic Acids Res. 25, 2816–2822 (1997).
    [CrossRef] [PubMed]
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    [CrossRef] [PubMed]
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    [CrossRef]
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    [CrossRef] [PubMed]
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    [CrossRef] [PubMed]
  13. C. Xu, J. B. Shear, W. W. Webb, “Hyper Rayleigh and hyper Raman scattering background of liquid water in two photon excited fluorescence detection,” Anal. Chem. 69, 1285–1287 (1997).
    [CrossRef] [PubMed]
  14. M. Fischer, C. D. Tran, “Thermal lens induced anomalous solvent effect on fluorescence produced by two photon continuous laser excitation,” Appl. Opt. 39, 6257–6262 (2000).
    [CrossRef]
  15. C. D. Tran, “Acousto-optic devices: optical elements for spectroscopy,” Anal. Chem. 64, 971A–981A (1992).
  16. C. D. Tran, J. Lu, “Characterization of the acousto-optic tunable filter for the ultraviolet and visible regions and development of an acousto-optic tunable filter based rapid scanning detector for high-performance liquid chromatography,” Anal. Chim. Acta 314, 57–66 (1995).
    [CrossRef]
  17. C. D. Tran, “Principles and analytical applications of acousto-optic tunable filters: an overview,” Talanta 45, 237–248 (1997).
    [CrossRef]
  18. C. D. Tran, R. J. Furlan, “Acousto-optic tunable filter as a polychromator and its application in multidimensional fluorescence spectrometry,” Anal. Chem. 64, 2775–2782 (1992).
    [CrossRef] [PubMed]
  19. C. D. Tran, R. J. Furlan, J. Lu, “Development of a multiwavelength thermal lens spectrophotometer based on an acousto-optic tunable filter as a polychromator,” Appl. Spectrosc. 48, 101–106 (1994).
    [CrossRef]
  20. C. D. Tran, R. J. Furlan, “Amplitude stabilization of a multiwavelength laser beam by an acousto-optic tunable filter,” Rev. Sci. Instrum. 65, 309–314 (1994).
    [CrossRef]

2000 (2)

H. He, L. B. McGown, “DNA sequencing by capillary electrophoresis with four-decay fluorescence detection,” Anal. Chem. 72, 5865–5873 (2000).
[CrossRef]

M. Fischer, C. D. Tran, “Thermal lens induced anomalous solvent effect on fluorescence produced by two photon continuous laser excitation,” Appl. Opt. 39, 6257–6262 (2000).
[CrossRef]

1998 (1)

H. He, B. K. Nunnally, L. Li, L. B. McGown, “On-the-fly fluorescence lifetime detection of dye labeled DNA primers for multiplex analysis,” Anal. Chem. 70, 3413–3418 (1998).
[CrossRef] [PubMed]

1997 (4)

L. G. Lee, S. L. Spergeon, C. R. Heiner, S. C. Benson, B. B. Resenblum, S. M. Menchen, R. J. Graham, A. Constantinescu, K. G. Upadhya, J. M. Cassel, “New energy transfer dyes for DNA sequencing,” Nucleic Acids Res. 25, 2816–2822 (1997).
[CrossRef] [PubMed]

S. M. Clark, R. A. Mathies, “Multiplex dsDNA fragment sizing using dimeric intercalation dyes and capillary array electrophoresis: ionic effects on the stability and electrophoretic mobility of DNA-dye complexes,” Anal. Chem. 69, 1355–1363 (1997).
[CrossRef] [PubMed]

C. D. Tran, “Principles and analytical applications of acousto-optic tunable filters: an overview,” Talanta 45, 237–248 (1997).
[CrossRef]

C. Xu, J. B. Shear, W. W. Webb, “Hyper Rayleigh and hyper Raman scattering background of liquid water in two photon excited fluorescence detection,” Anal. Chem. 69, 1285–1287 (1997).
[CrossRef] [PubMed]

1996 (1)

M. L. Metzker, J. Lu, R. A. Gibbs, “Electrophoretically uniform fluorescent dyes for automated DNA sequencing,” Science 271, 1420–1422 (1996).
[CrossRef] [PubMed]

1995 (3)

S. A. Soper, B. L. Legendre, D. C. Williams, “On-line fluorescence lifetime determinations in capillary electrophoresis,” Anal. Chem. 67, 4358–4365 (1995).
[CrossRef] [PubMed]

C. D. Tran, J. Lu, “Characterization of the acousto-optic tunable filter for the ultraviolet and visible regions and development of an acousto-optic tunable filter based rapid scanning detector for high-performance liquid chromatography,” Anal. Chim. Acta 314, 57–66 (1995).
[CrossRef]

A. Fischer, C. Cremer, E. H. K. Stelzer, “Fluorescence of coumarins and xanthenes after two-photon absorption with a pulsed titanium sapphire laser,” Appl. Opt. 34, 1989–2003 (1995).
[CrossRef] [PubMed]

1994 (2)

C. D. Tran, R. J. Furlan, J. Lu, “Development of a multiwavelength thermal lens spectrophotometer based on an acousto-optic tunable filter as a polychromator,” Appl. Spectrosc. 48, 101–106 (1994).
[CrossRef]

C. D. Tran, R. J. Furlan, “Amplitude stabilization of a multiwavelength laser beam by an acousto-optic tunable filter,” Rev. Sci. Instrum. 65, 309–314 (1994).
[CrossRef]

1993 (1)

L. M. Smith, “The future of DNA sequencing,” Science 262, 530–532 (1993).
[CrossRef] [PubMed]

1992 (2)

C. D. Tran, R. J. Furlan, “Acousto-optic tunable filter as a polychromator and its application in multidimensional fluorescence spectrometry,” Anal. Chem. 64, 2775–2782 (1992).
[CrossRef] [PubMed]

C. D. Tran, “Acousto-optic devices: optical elements for spectroscopy,” Anal. Chem. 64, 971A–981A (1992).

1990 (1)

W. Denk, J. H. Stricker, W. W. Webb, “Two photon laser scanning fluorescence microscopy,” Science 248, 73–76 (1990).
[CrossRef] [PubMed]

1986 (1)

L. M. Smith, J. Z. Sander, R. J. Kaiser, P. Hughes, C. Dodd, C. R. Connell, C. Heiner, S. B. H. Kent, L. E. Hood, “Fluorescence detection in automated DNA sequencing analysis,” Nature 321, 674–679 (1986).
[CrossRef] [PubMed]

1983 (1)

M. J. Wirth, A. C. Koskelo, C. E. Mohler, “Study of solvation symmetry by two photon polarization measurements,” J. Phys. Chem. 87, 4395–4400 (1983).
[CrossRef]

1931 (1)

M. Gopper-Mayer, “Uber Elementarakte mit zwei Quantensprungen,” Ann. Phys. (Leipzig) 9, 273–295 (1931).

Benson, S. C.

L. G. Lee, S. L. Spergeon, C. R. Heiner, S. C. Benson, B. B. Resenblum, S. M. Menchen, R. J. Graham, A. Constantinescu, K. G. Upadhya, J. M. Cassel, “New energy transfer dyes for DNA sequencing,” Nucleic Acids Res. 25, 2816–2822 (1997).
[CrossRef] [PubMed]

Cassel, J. M.

L. G. Lee, S. L. Spergeon, C. R. Heiner, S. C. Benson, B. B. Resenblum, S. M. Menchen, R. J. Graham, A. Constantinescu, K. G. Upadhya, J. M. Cassel, “New energy transfer dyes for DNA sequencing,” Nucleic Acids Res. 25, 2816–2822 (1997).
[CrossRef] [PubMed]

Clark, S. M.

S. M. Clark, R. A. Mathies, “Multiplex dsDNA fragment sizing using dimeric intercalation dyes and capillary array electrophoresis: ionic effects on the stability and electrophoretic mobility of DNA-dye complexes,” Anal. Chem. 69, 1355–1363 (1997).
[CrossRef] [PubMed]

Connell, C. R.

L. M. Smith, J. Z. Sander, R. J. Kaiser, P. Hughes, C. Dodd, C. R. Connell, C. Heiner, S. B. H. Kent, L. E. Hood, “Fluorescence detection in automated DNA sequencing analysis,” Nature 321, 674–679 (1986).
[CrossRef] [PubMed]

Constantinescu, A.

L. G. Lee, S. L. Spergeon, C. R. Heiner, S. C. Benson, B. B. Resenblum, S. M. Menchen, R. J. Graham, A. Constantinescu, K. G. Upadhya, J. M. Cassel, “New energy transfer dyes for DNA sequencing,” Nucleic Acids Res. 25, 2816–2822 (1997).
[CrossRef] [PubMed]

Cremer, C.

Denk, W.

W. Denk, J. H. Stricker, W. W. Webb, “Two photon laser scanning fluorescence microscopy,” Science 248, 73–76 (1990).
[CrossRef] [PubMed]

Dodd, C.

L. M. Smith, J. Z. Sander, R. J. Kaiser, P. Hughes, C. Dodd, C. R. Connell, C. Heiner, S. B. H. Kent, L. E. Hood, “Fluorescence detection in automated DNA sequencing analysis,” Nature 321, 674–679 (1986).
[CrossRef] [PubMed]

Fischer, A.

Fischer, M.

Furlan, R. J.

C. D. Tran, R. J. Furlan, “Amplitude stabilization of a multiwavelength laser beam by an acousto-optic tunable filter,” Rev. Sci. Instrum. 65, 309–314 (1994).
[CrossRef]

C. D. Tran, R. J. Furlan, J. Lu, “Development of a multiwavelength thermal lens spectrophotometer based on an acousto-optic tunable filter as a polychromator,” Appl. Spectrosc. 48, 101–106 (1994).
[CrossRef]

C. D. Tran, R. J. Furlan, “Acousto-optic tunable filter as a polychromator and its application in multidimensional fluorescence spectrometry,” Anal. Chem. 64, 2775–2782 (1992).
[CrossRef] [PubMed]

Gibbs, R. A.

M. L. Metzker, J. Lu, R. A. Gibbs, “Electrophoretically uniform fluorescent dyes for automated DNA sequencing,” Science 271, 1420–1422 (1996).
[CrossRef] [PubMed]

Gopper-Mayer, M.

M. Gopper-Mayer, “Uber Elementarakte mit zwei Quantensprungen,” Ann. Phys. (Leipzig) 9, 273–295 (1931).

Graham, R. J.

L. G. Lee, S. L. Spergeon, C. R. Heiner, S. C. Benson, B. B. Resenblum, S. M. Menchen, R. J. Graham, A. Constantinescu, K. G. Upadhya, J. M. Cassel, “New energy transfer dyes for DNA sequencing,” Nucleic Acids Res. 25, 2816–2822 (1997).
[CrossRef] [PubMed]

He, H.

H. He, L. B. McGown, “DNA sequencing by capillary electrophoresis with four-decay fluorescence detection,” Anal. Chem. 72, 5865–5873 (2000).
[CrossRef]

H. He, B. K. Nunnally, L. Li, L. B. McGown, “On-the-fly fluorescence lifetime detection of dye labeled DNA primers for multiplex analysis,” Anal. Chem. 70, 3413–3418 (1998).
[CrossRef] [PubMed]

Heiner, C.

L. M. Smith, J. Z. Sander, R. J. Kaiser, P. Hughes, C. Dodd, C. R. Connell, C. Heiner, S. B. H. Kent, L. E. Hood, “Fluorescence detection in automated DNA sequencing analysis,” Nature 321, 674–679 (1986).
[CrossRef] [PubMed]

Heiner, C. R.

L. G. Lee, S. L. Spergeon, C. R. Heiner, S. C. Benson, B. B. Resenblum, S. M. Menchen, R. J. Graham, A. Constantinescu, K. G. Upadhya, J. M. Cassel, “New energy transfer dyes for DNA sequencing,” Nucleic Acids Res. 25, 2816–2822 (1997).
[CrossRef] [PubMed]

Hood, L. E.

L. M. Smith, J. Z. Sander, R. J. Kaiser, P. Hughes, C. Dodd, C. R. Connell, C. Heiner, S. B. H. Kent, L. E. Hood, “Fluorescence detection in automated DNA sequencing analysis,” Nature 321, 674–679 (1986).
[CrossRef] [PubMed]

Hughes, P.

L. M. Smith, J. Z. Sander, R. J. Kaiser, P. Hughes, C. Dodd, C. R. Connell, C. Heiner, S. B. H. Kent, L. E. Hood, “Fluorescence detection in automated DNA sequencing analysis,” Nature 321, 674–679 (1986).
[CrossRef] [PubMed]

Kaiser, R. J.

L. M. Smith, J. Z. Sander, R. J. Kaiser, P. Hughes, C. Dodd, C. R. Connell, C. Heiner, S. B. H. Kent, L. E. Hood, “Fluorescence detection in automated DNA sequencing analysis,” Nature 321, 674–679 (1986).
[CrossRef] [PubMed]

Kent, S. B. H.

L. M. Smith, J. Z. Sander, R. J. Kaiser, P. Hughes, C. Dodd, C. R. Connell, C. Heiner, S. B. H. Kent, L. E. Hood, “Fluorescence detection in automated DNA sequencing analysis,” Nature 321, 674–679 (1986).
[CrossRef] [PubMed]

Koskelo, A. C.

M. J. Wirth, A. C. Koskelo, C. E. Mohler, “Study of solvation symmetry by two photon polarization measurements,” J. Phys. Chem. 87, 4395–4400 (1983).
[CrossRef]

Lee, L. G.

L. G. Lee, S. L. Spergeon, C. R. Heiner, S. C. Benson, B. B. Resenblum, S. M. Menchen, R. J. Graham, A. Constantinescu, K. G. Upadhya, J. M. Cassel, “New energy transfer dyes for DNA sequencing,” Nucleic Acids Res. 25, 2816–2822 (1997).
[CrossRef] [PubMed]

Legendre, B. L.

S. A. Soper, B. L. Legendre, D. C. Williams, “On-line fluorescence lifetime determinations in capillary electrophoresis,” Anal. Chem. 67, 4358–4365 (1995).
[CrossRef] [PubMed]

Li, L.

H. He, B. K. Nunnally, L. Li, L. B. McGown, “On-the-fly fluorescence lifetime detection of dye labeled DNA primers for multiplex analysis,” Anal. Chem. 70, 3413–3418 (1998).
[CrossRef] [PubMed]

Lu, J.

M. L. Metzker, J. Lu, R. A. Gibbs, “Electrophoretically uniform fluorescent dyes for automated DNA sequencing,” Science 271, 1420–1422 (1996).
[CrossRef] [PubMed]

C. D. Tran, J. Lu, “Characterization of the acousto-optic tunable filter for the ultraviolet and visible regions and development of an acousto-optic tunable filter based rapid scanning detector for high-performance liquid chromatography,” Anal. Chim. Acta 314, 57–66 (1995).
[CrossRef]

C. D. Tran, R. J. Furlan, J. Lu, “Development of a multiwavelength thermal lens spectrophotometer based on an acousto-optic tunable filter as a polychromator,” Appl. Spectrosc. 48, 101–106 (1994).
[CrossRef]

Mathies, R. A.

S. M. Clark, R. A. Mathies, “Multiplex dsDNA fragment sizing using dimeric intercalation dyes and capillary array electrophoresis: ionic effects on the stability and electrophoretic mobility of DNA-dye complexes,” Anal. Chem. 69, 1355–1363 (1997).
[CrossRef] [PubMed]

McGown, L. B.

H. He, L. B. McGown, “DNA sequencing by capillary electrophoresis with four-decay fluorescence detection,” Anal. Chem. 72, 5865–5873 (2000).
[CrossRef]

H. He, B. K. Nunnally, L. Li, L. B. McGown, “On-the-fly fluorescence lifetime detection of dye labeled DNA primers for multiplex analysis,” Anal. Chem. 70, 3413–3418 (1998).
[CrossRef] [PubMed]

Menchen, S. M.

L. G. Lee, S. L. Spergeon, C. R. Heiner, S. C. Benson, B. B. Resenblum, S. M. Menchen, R. J. Graham, A. Constantinescu, K. G. Upadhya, J. M. Cassel, “New energy transfer dyes for DNA sequencing,” Nucleic Acids Res. 25, 2816–2822 (1997).
[CrossRef] [PubMed]

Metzker, M. L.

M. L. Metzker, J. Lu, R. A. Gibbs, “Electrophoretically uniform fluorescent dyes for automated DNA sequencing,” Science 271, 1420–1422 (1996).
[CrossRef] [PubMed]

Mohler, C. E.

M. J. Wirth, A. C. Koskelo, C. E. Mohler, “Study of solvation symmetry by two photon polarization measurements,” J. Phys. Chem. 87, 4395–4400 (1983).
[CrossRef]

Nunnally, B. K.

H. He, B. K. Nunnally, L. Li, L. B. McGown, “On-the-fly fluorescence lifetime detection of dye labeled DNA primers for multiplex analysis,” Anal. Chem. 70, 3413–3418 (1998).
[CrossRef] [PubMed]

Resenblum, B. B.

L. G. Lee, S. L. Spergeon, C. R. Heiner, S. C. Benson, B. B. Resenblum, S. M. Menchen, R. J. Graham, A. Constantinescu, K. G. Upadhya, J. M. Cassel, “New energy transfer dyes for DNA sequencing,” Nucleic Acids Res. 25, 2816–2822 (1997).
[CrossRef] [PubMed]

Sander, J. Z.

L. M. Smith, J. Z. Sander, R. J. Kaiser, P. Hughes, C. Dodd, C. R. Connell, C. Heiner, S. B. H. Kent, L. E. Hood, “Fluorescence detection in automated DNA sequencing analysis,” Nature 321, 674–679 (1986).
[CrossRef] [PubMed]

Shear, J. B.

C. Xu, J. B. Shear, W. W. Webb, “Hyper Rayleigh and hyper Raman scattering background of liquid water in two photon excited fluorescence detection,” Anal. Chem. 69, 1285–1287 (1997).
[CrossRef] [PubMed]

Smith, L. M.

L. M. Smith, “The future of DNA sequencing,” Science 262, 530–532 (1993).
[CrossRef] [PubMed]

L. M. Smith, J. Z. Sander, R. J. Kaiser, P. Hughes, C. Dodd, C. R. Connell, C. Heiner, S. B. H. Kent, L. E. Hood, “Fluorescence detection in automated DNA sequencing analysis,” Nature 321, 674–679 (1986).
[CrossRef] [PubMed]

Soper, S. A.

S. A. Soper, B. L. Legendre, D. C. Williams, “On-line fluorescence lifetime determinations in capillary electrophoresis,” Anal. Chem. 67, 4358–4365 (1995).
[CrossRef] [PubMed]

Spergeon, S. L.

L. G. Lee, S. L. Spergeon, C. R. Heiner, S. C. Benson, B. B. Resenblum, S. M. Menchen, R. J. Graham, A. Constantinescu, K. G. Upadhya, J. M. Cassel, “New energy transfer dyes for DNA sequencing,” Nucleic Acids Res. 25, 2816–2822 (1997).
[CrossRef] [PubMed]

Stelzer, E. H. K.

Stricker, J. H.

W. Denk, J. H. Stricker, W. W. Webb, “Two photon laser scanning fluorescence microscopy,” Science 248, 73–76 (1990).
[CrossRef] [PubMed]

Tran, C. D.

M. Fischer, C. D. Tran, “Thermal lens induced anomalous solvent effect on fluorescence produced by two photon continuous laser excitation,” Appl. Opt. 39, 6257–6262 (2000).
[CrossRef]

C. D. Tran, “Principles and analytical applications of acousto-optic tunable filters: an overview,” Talanta 45, 237–248 (1997).
[CrossRef]

C. D. Tran, J. Lu, “Characterization of the acousto-optic tunable filter for the ultraviolet and visible regions and development of an acousto-optic tunable filter based rapid scanning detector for high-performance liquid chromatography,” Anal. Chim. Acta 314, 57–66 (1995).
[CrossRef]

C. D. Tran, R. J. Furlan, “Amplitude stabilization of a multiwavelength laser beam by an acousto-optic tunable filter,” Rev. Sci. Instrum. 65, 309–314 (1994).
[CrossRef]

C. D. Tran, R. J. Furlan, J. Lu, “Development of a multiwavelength thermal lens spectrophotometer based on an acousto-optic tunable filter as a polychromator,” Appl. Spectrosc. 48, 101–106 (1994).
[CrossRef]

C. D. Tran, “Acousto-optic devices: optical elements for spectroscopy,” Anal. Chem. 64, 971A–981A (1992).

C. D. Tran, R. J. Furlan, “Acousto-optic tunable filter as a polychromator and its application in multidimensional fluorescence spectrometry,” Anal. Chem. 64, 2775–2782 (1992).
[CrossRef] [PubMed]

Upadhya, K. G.

L. G. Lee, S. L. Spergeon, C. R. Heiner, S. C. Benson, B. B. Resenblum, S. M. Menchen, R. J. Graham, A. Constantinescu, K. G. Upadhya, J. M. Cassel, “New energy transfer dyes for DNA sequencing,” Nucleic Acids Res. 25, 2816–2822 (1997).
[CrossRef] [PubMed]

Webb, W. W.

C. Xu, J. B. Shear, W. W. Webb, “Hyper Rayleigh and hyper Raman scattering background of liquid water in two photon excited fluorescence detection,” Anal. Chem. 69, 1285–1287 (1997).
[CrossRef] [PubMed]

W. Denk, J. H. Stricker, W. W. Webb, “Two photon laser scanning fluorescence microscopy,” Science 248, 73–76 (1990).
[CrossRef] [PubMed]

Williams, D. C.

S. A. Soper, B. L. Legendre, D. C. Williams, “On-line fluorescence lifetime determinations in capillary electrophoresis,” Anal. Chem. 67, 4358–4365 (1995).
[CrossRef] [PubMed]

Wirth, M. J.

M. J. Wirth, A. C. Koskelo, C. E. Mohler, “Study of solvation symmetry by two photon polarization measurements,” J. Phys. Chem. 87, 4395–4400 (1983).
[CrossRef]

Xu, C.

C. Xu, J. B. Shear, W. W. Webb, “Hyper Rayleigh and hyper Raman scattering background of liquid water in two photon excited fluorescence detection,” Anal. Chem. 69, 1285–1287 (1997).
[CrossRef] [PubMed]

Anal. Chem. (7)

S. M. Clark, R. A. Mathies, “Multiplex dsDNA fragment sizing using dimeric intercalation dyes and capillary array electrophoresis: ionic effects on the stability and electrophoretic mobility of DNA-dye complexes,” Anal. Chem. 69, 1355–1363 (1997).
[CrossRef] [PubMed]

S. A. Soper, B. L. Legendre, D. C. Williams, “On-line fluorescence lifetime determinations in capillary electrophoresis,” Anal. Chem. 67, 4358–4365 (1995).
[CrossRef] [PubMed]

H. He, L. B. McGown, “DNA sequencing by capillary electrophoresis with four-decay fluorescence detection,” Anal. Chem. 72, 5865–5873 (2000).
[CrossRef]

H. He, B. K. Nunnally, L. Li, L. B. McGown, “On-the-fly fluorescence lifetime detection of dye labeled DNA primers for multiplex analysis,” Anal. Chem. 70, 3413–3418 (1998).
[CrossRef] [PubMed]

C. Xu, J. B. Shear, W. W. Webb, “Hyper Rayleigh and hyper Raman scattering background of liquid water in two photon excited fluorescence detection,” Anal. Chem. 69, 1285–1287 (1997).
[CrossRef] [PubMed]

C. D. Tran, “Acousto-optic devices: optical elements for spectroscopy,” Anal. Chem. 64, 971A–981A (1992).

C. D. Tran, R. J. Furlan, “Acousto-optic tunable filter as a polychromator and its application in multidimensional fluorescence spectrometry,” Anal. Chem. 64, 2775–2782 (1992).
[CrossRef] [PubMed]

Anal. Chim. Acta (1)

C. D. Tran, J. Lu, “Characterization of the acousto-optic tunable filter for the ultraviolet and visible regions and development of an acousto-optic tunable filter based rapid scanning detector for high-performance liquid chromatography,” Anal. Chim. Acta 314, 57–66 (1995).
[CrossRef]

Ann. Phys. (Leipzig) (1)

M. Gopper-Mayer, “Uber Elementarakte mit zwei Quantensprungen,” Ann. Phys. (Leipzig) 9, 273–295 (1931).

Appl. Opt. (2)

Appl. Spectrosc. (1)

J. Phys. Chem. (1)

M. J. Wirth, A. C. Koskelo, C. E. Mohler, “Study of solvation symmetry by two photon polarization measurements,” J. Phys. Chem. 87, 4395–4400 (1983).
[CrossRef]

Nature (1)

L. M. Smith, J. Z. Sander, R. J. Kaiser, P. Hughes, C. Dodd, C. R. Connell, C. Heiner, S. B. H. Kent, L. E. Hood, “Fluorescence detection in automated DNA sequencing analysis,” Nature 321, 674–679 (1986).
[CrossRef] [PubMed]

Nucleic Acids Res. (1)

L. G. Lee, S. L. Spergeon, C. R. Heiner, S. C. Benson, B. B. Resenblum, S. M. Menchen, R. J. Graham, A. Constantinescu, K. G. Upadhya, J. M. Cassel, “New energy transfer dyes for DNA sequencing,” Nucleic Acids Res. 25, 2816–2822 (1997).
[CrossRef] [PubMed]

Rev. Sci. Instrum. (1)

C. D. Tran, R. J. Furlan, “Amplitude stabilization of a multiwavelength laser beam by an acousto-optic tunable filter,” Rev. Sci. Instrum. 65, 309–314 (1994).
[CrossRef]

Science (3)

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[CrossRef]

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Figures (7)

Fig. 1
Fig. 1

Schematic diagram of the AOTF-based fluorimeter: AOTF, acousto-optic tunable filter; lock-in, lock-in amplifier; PMT, photomultiplier tube; A/D, analog to digital converter; D/A, digital to analog converter.

Fig. 2
Fig. 2

One-photon fluorescence spectrum of 2.85 × 10-6 M ADS 840NCS uncoupled dye in 6.25 × 10-2 M SDS (excitation wavelength, 845 nm).

Fig. 3
Fig. 3

One-photon-excited fluorescence spectrum of Rhodamine Red-X in 6.25 × 10-2 M, obtained with 520-nm excitation light.

Fig. 4
Fig. 4

Two-photon excitation spectrum of Rhodamine Red-X in 6.25 × 10-2M (monitored at 606 nm).

Fig. 5
Fig. 5

Two-photon-excited fluorescence spectrum of 1.0 × 10-5 M Rhodamine Red-X in 6.25 × 10-2 M SDS, excited at 845 nm.

Fig. 6
Fig. 6

Plot of intensity of two-photon-excited fluorescence versus excitation laser power for 1.0 × 10-5 M Rhodamine Red-X.

Fig. 7
Fig. 7

One- and two-photon-excited fluorescence spectra of a sample containing ADS 840NCS dye coupled to an oligonucleotide, and Rhodamine Red-X coupled to an oligonucleotide. The spectra were obtained through an excitation of the sample with a single excitation wavelength of 845 nm; the dashed line is the one-photon fluorescence spectrum of the ADS 840NCS coupled oligonucleotide and the solid line is the two-photon excited fluorescence spectrum of Rhodamine Red-X coupled oligonucleotide.

Tables (1)

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Table 1 Oligonucleotide Labeling Procedure

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