Abstract

We describe a novel technique for DNA-microarray reading based on time-resolved fluorescence measurements. We used an intensified CCD camera with picosecond resolution to acquire a set of time-delayed fluorescence images from a mutation DNA microarray marked with cyanine 3. We measured the fluorescence lifetimes of the marker and the background separately, and we used this information to calculate the amplitude map of the marker, starting from the time-delayed images. This procedure allowed us to identify hybridized spots that are not visible in fluorescence images acquired with continuous-wave detection.

© 2000 Optical Society of America

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References

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  1. M. S. Chee, R. Yang, E. Hubbell, A. Berno, X. C. Huang, D. Stern, J. Winkler, D. J. Lockhart, M. S. Morris, and S. P. A. Fodor, Science 274, 610 (1996).
    [CrossRef] [PubMed]
  2. J. Khan, L. Bittner, P. S. Meltzer, and J. M. Trent, Biochim. Biophys. Acta 1423, M17 (1999).
    [PubMed]
  3. E. S. Lander, Nature Genetics 21, 3 (1999).
    [CrossRef]
  4. L. Wodicka, H. Dong, M. Mittmann, M.-H. Ho, and D. J. Lockhart, Nature Biotechnol. 15, 1359 (1997).
    [CrossRef]
  5. C. A. Afshary, E. F. Nuwaysir, and J. C. Barrett, Cancer Res. 59, 4759 (1999).
  6. R. Cubeddu, A. Pifferi, P. Taroni, and G. Valentini, Opt. Lett. 20, 2553 (1995).
    [CrossRef]
  7. K. Dowling, M. J. Dayel, M. J. Lever, P. M. W. French, J. D. Hares, and A. K. L. Dymoke-Bradshaw, Opt. Lett. 23, 810 (1998).
    [CrossRef]

1999 (3)

J. Khan, L. Bittner, P. S. Meltzer, and J. M. Trent, Biochim. Biophys. Acta 1423, M17 (1999).
[PubMed]

E. S. Lander, Nature Genetics 21, 3 (1999).
[CrossRef]

C. A. Afshary, E. F. Nuwaysir, and J. C. Barrett, Cancer Res. 59, 4759 (1999).

1998 (1)

1997 (1)

L. Wodicka, H. Dong, M. Mittmann, M.-H. Ho, and D. J. Lockhart, Nature Biotechnol. 15, 1359 (1997).
[CrossRef]

1996 (1)

M. S. Chee, R. Yang, E. Hubbell, A. Berno, X. C. Huang, D. Stern, J. Winkler, D. J. Lockhart, M. S. Morris, and S. P. A. Fodor, Science 274, 610 (1996).
[CrossRef] [PubMed]

1995 (1)

Afshary, C. A.

C. A. Afshary, E. F. Nuwaysir, and J. C. Barrett, Cancer Res. 59, 4759 (1999).

Barrett, J. C.

C. A. Afshary, E. F. Nuwaysir, and J. C. Barrett, Cancer Res. 59, 4759 (1999).

Berno, A.

M. S. Chee, R. Yang, E. Hubbell, A. Berno, X. C. Huang, D. Stern, J. Winkler, D. J. Lockhart, M. S. Morris, and S. P. A. Fodor, Science 274, 610 (1996).
[CrossRef] [PubMed]

Bittner, L.

J. Khan, L. Bittner, P. S. Meltzer, and J. M. Trent, Biochim. Biophys. Acta 1423, M17 (1999).
[PubMed]

Chee, M. S.

M. S. Chee, R. Yang, E. Hubbell, A. Berno, X. C. Huang, D. Stern, J. Winkler, D. J. Lockhart, M. S. Morris, and S. P. A. Fodor, Science 274, 610 (1996).
[CrossRef] [PubMed]

Cubeddu, R.

Dayel, M. J.

Dong, H.

L. Wodicka, H. Dong, M. Mittmann, M.-H. Ho, and D. J. Lockhart, Nature Biotechnol. 15, 1359 (1997).
[CrossRef]

Dowling, K.

Dymoke-Bradshaw, A. K. L.

Fodor, S. P. A.

M. S. Chee, R. Yang, E. Hubbell, A. Berno, X. C. Huang, D. Stern, J. Winkler, D. J. Lockhart, M. S. Morris, and S. P. A. Fodor, Science 274, 610 (1996).
[CrossRef] [PubMed]

French, P. M. W.

Hares, J. D.

Ho, M.-H.

L. Wodicka, H. Dong, M. Mittmann, M.-H. Ho, and D. J. Lockhart, Nature Biotechnol. 15, 1359 (1997).
[CrossRef]

Huang, X. C.

M. S. Chee, R. Yang, E. Hubbell, A. Berno, X. C. Huang, D. Stern, J. Winkler, D. J. Lockhart, M. S. Morris, and S. P. A. Fodor, Science 274, 610 (1996).
[CrossRef] [PubMed]

Hubbell, E.

M. S. Chee, R. Yang, E. Hubbell, A. Berno, X. C. Huang, D. Stern, J. Winkler, D. J. Lockhart, M. S. Morris, and S. P. A. Fodor, Science 274, 610 (1996).
[CrossRef] [PubMed]

Khan, J.

J. Khan, L. Bittner, P. S. Meltzer, and J. M. Trent, Biochim. Biophys. Acta 1423, M17 (1999).
[PubMed]

Lander, E. S.

E. S. Lander, Nature Genetics 21, 3 (1999).
[CrossRef]

Lever, M. J.

Lockhart, D. J.

L. Wodicka, H. Dong, M. Mittmann, M.-H. Ho, and D. J. Lockhart, Nature Biotechnol. 15, 1359 (1997).
[CrossRef]

M. S. Chee, R. Yang, E. Hubbell, A. Berno, X. C. Huang, D. Stern, J. Winkler, D. J. Lockhart, M. S. Morris, and S. P. A. Fodor, Science 274, 610 (1996).
[CrossRef] [PubMed]

Meltzer, P. S.

J. Khan, L. Bittner, P. S. Meltzer, and J. M. Trent, Biochim. Biophys. Acta 1423, M17 (1999).
[PubMed]

Mittmann, M.

L. Wodicka, H. Dong, M. Mittmann, M.-H. Ho, and D. J. Lockhart, Nature Biotechnol. 15, 1359 (1997).
[CrossRef]

Morris, M. S.

M. S. Chee, R. Yang, E. Hubbell, A. Berno, X. C. Huang, D. Stern, J. Winkler, D. J. Lockhart, M. S. Morris, and S. P. A. Fodor, Science 274, 610 (1996).
[CrossRef] [PubMed]

Nuwaysir, E. F.

C. A. Afshary, E. F. Nuwaysir, and J. C. Barrett, Cancer Res. 59, 4759 (1999).

Pifferi, A.

Stern, D.

M. S. Chee, R. Yang, E. Hubbell, A. Berno, X. C. Huang, D. Stern, J. Winkler, D. J. Lockhart, M. S. Morris, and S. P. A. Fodor, Science 274, 610 (1996).
[CrossRef] [PubMed]

Taroni, P.

Trent, J. M.

J. Khan, L. Bittner, P. S. Meltzer, and J. M. Trent, Biochim. Biophys. Acta 1423, M17 (1999).
[PubMed]

Valentini, G.

Winkler, J.

M. S. Chee, R. Yang, E. Hubbell, A. Berno, X. C. Huang, D. Stern, J. Winkler, D. J. Lockhart, M. S. Morris, and S. P. A. Fodor, Science 274, 610 (1996).
[CrossRef] [PubMed]

Wodicka, L.

L. Wodicka, H. Dong, M. Mittmann, M.-H. Ho, and D. J. Lockhart, Nature Biotechnol. 15, 1359 (1997).
[CrossRef]

Yang, R.

M. S. Chee, R. Yang, E. Hubbell, A. Berno, X. C. Huang, D. Stern, J. Winkler, D. J. Lockhart, M. S. Morris, and S. P. A. Fodor, Science 274, 610 (1996).
[CrossRef] [PubMed]

Biochim. Biophys. Acta (1)

J. Khan, L. Bittner, P. S. Meltzer, and J. M. Trent, Biochim. Biophys. Acta 1423, M17 (1999).
[PubMed]

Cancer Res. (1)

C. A. Afshary, E. F. Nuwaysir, and J. C. Barrett, Cancer Res. 59, 4759 (1999).

Nature Biotechnol. (1)

L. Wodicka, H. Dong, M. Mittmann, M.-H. Ho, and D. J. Lockhart, Nature Biotechnol. 15, 1359 (1997).
[CrossRef]

Nature Genetics (1)

E. S. Lander, Nature Genetics 21, 3 (1999).
[CrossRef]

Opt. Lett. (2)

Science (1)

M. S. Chee, R. Yang, E. Hubbell, A. Berno, X. C. Huang, D. Stern, J. Winkler, D. J. Lockhart, M. S. Morris, and S. P. A. Fodor, Science 274, 610 (1996).
[CrossRef] [PubMed]

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Figures (4)

Fig. 1
Fig. 1

Schematic diagram of the experimental setup: BS, beam splitter; Pd, photodiode; CFD, constant-fraction discriminator; D, delay unit. ML, mode locked.

Fig. 2
Fig. 2

Fluorescence image of a DNA-microarray labeled with Cy3 (0.5 nM). The image was taken synchronously with the excitation pulses. The line profile across the first column is shown at the left.

Fig. 3
Fig. 3

Map of the amplitude of the Cy3 fluorescences obtained from a bilinear fit carried out on the set of time-delayed images of the DNA microarray considered in Fig. 2. The line profile across the first column is shown at the left.

Fig. 4
Fig. 4

Map of the amplitude of the background fluorescence obtained from a bilinear fit carried out on the set of time-delayed images of the DNA microarray considered in Fig. 2. The line profile across the first column is shown at the left.

Tables (1)

Tables Icon

Table 1 Average Fluorescence Lifetimes τ¯ and Normalized Amplitudes A¯ with Standard Deviations σ of Two Dyes and Their Mixture Measured with the Time-Resolved Imaging Systema

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