Abstract

The long (but not too long) fluorescence lifetime of CdSe semiconductor quantum dots was exploited to enhance fluorescence biological imaging contrast and sensitivity by time-gated detection. Significant and selective reduction of the autofluorescence contribution to the overall image was achieved, and enhancement of the signal-to-background ratio by more than an order of magnitude was demonstrated.

© 2001 Optical Society of America

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  1. M. J. Bruchez, M. M. Moronne, P. Gin, S. Weiss, and A. P. Alivisatos, Science 281, 2013 (1998).
    [CrossRef] [PubMed]
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    [CrossRef] [PubMed]
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    [CrossRef] [PubMed]
  4. E. Terpetschnig, H. Szmacinski, H. Malak, and J. R. Lakowicz, Biophys. J. 68, 342 (1995).
    [CrossRef] [PubMed]
  5. M. Li and P. R. Selvin, J. Am. Chem. Soc. 117, 8132 (1995).
    [CrossRef]
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    [CrossRef]
  7. J. R. Lakowicz, Scanning Microsc. Suppl.213 (1996).
  8. P. Tinnefeld, V. Buschmann, D.-P. Herten, K.-T. Han, and M. Sauer, Single Mol. 1, 3 215 (2000).
    [CrossRef]
  9. T. D. Lacoste, X. Michalet, F. Pinaud, D. S. Chemla, A. P. Alivisatos, and S. Weiss, Proc. Natl. Acad. Sci. USA 97, 9461 (2000).
    [CrossRef]

2000 (2)

P. Tinnefeld, V. Buschmann, D.-P. Herten, K.-T. Han, and M. Sauer, Single Mol. 1, 3 215 (2000).
[CrossRef]

T. D. Lacoste, X. Michalet, F. Pinaud, D. S. Chemla, A. P. Alivisatos, and S. Weiss, Proc. Natl. Acad. Sci. USA 97, 9461 (2000).
[CrossRef]

1998 (2)

M. J. Bruchez, M. M. Moronne, P. Gin, S. Weiss, and A. P. Alivisatos, Science 281, 2013 (1998).
[CrossRef] [PubMed]

W. C. W. Chan and S. Nie, Science 281, 2016 (1998).
[CrossRef] [PubMed]

1996 (2)

P. R. Selvin, J. Jancarik, M. Li, and L. W. Hung, Inorg. Chem. 35, 700 (1996).
[CrossRef]

J. R. Lakowicz, Scanning Microsc. Suppl.213 (1996).

1995 (2)

E. Terpetschnig, H. Szmacinski, H. Malak, and J. R. Lakowicz, Biophys. J. 68, 342 (1995).
[CrossRef] [PubMed]

M. Li and P. R. Selvin, J. Am. Chem. Soc. 117, 8132 (1995).
[CrossRef]

1991 (1)

G. Marriott, R. M. Clegg, D. J. Arndt-Jovin, and T. M. Jovin, Biophys. J. 60, 1374 (1991).
[CrossRef] [PubMed]

Alivisatos, A. P.

T. D. Lacoste, X. Michalet, F. Pinaud, D. S. Chemla, A. P. Alivisatos, and S. Weiss, Proc. Natl. Acad. Sci. USA 97, 9461 (2000).
[CrossRef]

M. J. Bruchez, M. M. Moronne, P. Gin, S. Weiss, and A. P. Alivisatos, Science 281, 2013 (1998).
[CrossRef] [PubMed]

Arndt-Jovin, D. J.

G. Marriott, R. M. Clegg, D. J. Arndt-Jovin, and T. M. Jovin, Biophys. J. 60, 1374 (1991).
[CrossRef] [PubMed]

Bruchez, M. J.

M. J. Bruchez, M. M. Moronne, P. Gin, S. Weiss, and A. P. Alivisatos, Science 281, 2013 (1998).
[CrossRef] [PubMed]

Buschmann, V.

P. Tinnefeld, V. Buschmann, D.-P. Herten, K.-T. Han, and M. Sauer, Single Mol. 1, 3 215 (2000).
[CrossRef]

Chan, W. C. W.

W. C. W. Chan and S. Nie, Science 281, 2016 (1998).
[CrossRef] [PubMed]

Chemla, D. S.

T. D. Lacoste, X. Michalet, F. Pinaud, D. S. Chemla, A. P. Alivisatos, and S. Weiss, Proc. Natl. Acad. Sci. USA 97, 9461 (2000).
[CrossRef]

Clegg, R. M.

G. Marriott, R. M. Clegg, D. J. Arndt-Jovin, and T. M. Jovin, Biophys. J. 60, 1374 (1991).
[CrossRef] [PubMed]

Gin, P.

M. J. Bruchez, M. M. Moronne, P. Gin, S. Weiss, and A. P. Alivisatos, Science 281, 2013 (1998).
[CrossRef] [PubMed]

Han, K.-T.

P. Tinnefeld, V. Buschmann, D.-P. Herten, K.-T. Han, and M. Sauer, Single Mol. 1, 3 215 (2000).
[CrossRef]

Herten, D.-P.

P. Tinnefeld, V. Buschmann, D.-P. Herten, K.-T. Han, and M. Sauer, Single Mol. 1, 3 215 (2000).
[CrossRef]

Hung, L. W.

P. R. Selvin, J. Jancarik, M. Li, and L. W. Hung, Inorg. Chem. 35, 700 (1996).
[CrossRef]

Jancarik, J.

P. R. Selvin, J. Jancarik, M. Li, and L. W. Hung, Inorg. Chem. 35, 700 (1996).
[CrossRef]

Jovin, T. M.

G. Marriott, R. M. Clegg, D. J. Arndt-Jovin, and T. M. Jovin, Biophys. J. 60, 1374 (1991).
[CrossRef] [PubMed]

Lacoste, T. D.

T. D. Lacoste, X. Michalet, F. Pinaud, D. S. Chemla, A. P. Alivisatos, and S. Weiss, Proc. Natl. Acad. Sci. USA 97, 9461 (2000).
[CrossRef]

Lakowicz, J. R.

J. R. Lakowicz, Scanning Microsc. Suppl.213 (1996).

E. Terpetschnig, H. Szmacinski, H. Malak, and J. R. Lakowicz, Biophys. J. 68, 342 (1995).
[CrossRef] [PubMed]

Li, M.

P. R. Selvin, J. Jancarik, M. Li, and L. W. Hung, Inorg. Chem. 35, 700 (1996).
[CrossRef]

M. Li and P. R. Selvin, J. Am. Chem. Soc. 117, 8132 (1995).
[CrossRef]

Malak, H.

E. Terpetschnig, H. Szmacinski, H. Malak, and J. R. Lakowicz, Biophys. J. 68, 342 (1995).
[CrossRef] [PubMed]

Marriott, G.

G. Marriott, R. M. Clegg, D. J. Arndt-Jovin, and T. M. Jovin, Biophys. J. 60, 1374 (1991).
[CrossRef] [PubMed]

Michalet, X.

T. D. Lacoste, X. Michalet, F. Pinaud, D. S. Chemla, A. P. Alivisatos, and S. Weiss, Proc. Natl. Acad. Sci. USA 97, 9461 (2000).
[CrossRef]

Moronne, M. M.

M. J. Bruchez, M. M. Moronne, P. Gin, S. Weiss, and A. P. Alivisatos, Science 281, 2013 (1998).
[CrossRef] [PubMed]

Nie, S.

W. C. W. Chan and S. Nie, Science 281, 2016 (1998).
[CrossRef] [PubMed]

Pinaud, F.

T. D. Lacoste, X. Michalet, F. Pinaud, D. S. Chemla, A. P. Alivisatos, and S. Weiss, Proc. Natl. Acad. Sci. USA 97, 9461 (2000).
[CrossRef]

Sauer, M.

P. Tinnefeld, V. Buschmann, D.-P. Herten, K.-T. Han, and M. Sauer, Single Mol. 1, 3 215 (2000).
[CrossRef]

Selvin, P. R.

P. R. Selvin, J. Jancarik, M. Li, and L. W. Hung, Inorg. Chem. 35, 700 (1996).
[CrossRef]

M. Li and P. R. Selvin, J. Am. Chem. Soc. 117, 8132 (1995).
[CrossRef]

Szmacinski, H.

E. Terpetschnig, H. Szmacinski, H. Malak, and J. R. Lakowicz, Biophys. J. 68, 342 (1995).
[CrossRef] [PubMed]

Terpetschnig, E.

E. Terpetschnig, H. Szmacinski, H. Malak, and J. R. Lakowicz, Biophys. J. 68, 342 (1995).
[CrossRef] [PubMed]

Tinnefeld, P.

P. Tinnefeld, V. Buschmann, D.-P. Herten, K.-T. Han, and M. Sauer, Single Mol. 1, 3 215 (2000).
[CrossRef]

Weiss, S.

T. D. Lacoste, X. Michalet, F. Pinaud, D. S. Chemla, A. P. Alivisatos, and S. Weiss, Proc. Natl. Acad. Sci. USA 97, 9461 (2000).
[CrossRef]

M. J. Bruchez, M. M. Moronne, P. Gin, S. Weiss, and A. P. Alivisatos, Science 281, 2013 (1998).
[CrossRef] [PubMed]

Biophys. J. (2)

G. Marriott, R. M. Clegg, D. J. Arndt-Jovin, and T. M. Jovin, Biophys. J. 60, 1374 (1991).
[CrossRef] [PubMed]

E. Terpetschnig, H. Szmacinski, H. Malak, and J. R. Lakowicz, Biophys. J. 68, 342 (1995).
[CrossRef] [PubMed]

Inorg. Chem. (1)

P. R. Selvin, J. Jancarik, M. Li, and L. W. Hung, Inorg. Chem. 35, 700 (1996).
[CrossRef]

J. Am. Chem. Soc. (1)

M. Li and P. R. Selvin, J. Am. Chem. Soc. 117, 8132 (1995).
[CrossRef]

Proc. Natl. Acad. Sci. USA (1)

T. D. Lacoste, X. Michalet, F. Pinaud, D. S. Chemla, A. P. Alivisatos, and S. Weiss, Proc. Natl. Acad. Sci. USA 97, 9461 (2000).
[CrossRef]

Scanning Microsc. Suppl. (1)

J. R. Lakowicz, Scanning Microsc. Suppl.213 (1996).

Science (2)

M. J. Bruchez, M. M. Moronne, P. Gin, S. Weiss, and A. P. Alivisatos, Science 281, 2013 (1998).
[CrossRef] [PubMed]

W. C. W. Chan and S. Nie, Science 281, 2016 (1998).
[CrossRef] [PubMed]

Single Mol. (1)

P. Tinnefeld, V. Buschmann, D.-P. Herten, K.-T. Han, and M. Sauer, Single Mol. 1, 3 215 (2000).
[CrossRef]

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Figures (3)

Fig. 1
Fig. 1

Experimental setup: The microscope is a homemade stage-scanning confocal microscope with a nanometer-resolution closed-loop piezo-stage scanner. A single laser line is brought via a fiber and a beam expander to the back focal plane of the objective (Ob) after reflection on a dichroic mirror (DC). Fluorescence is collected by the same objective, filtered by a bandpass filter (BP), and focused onto a pinhole (PH) via a tube lens (TL). The pinhole is imaged onto an avalanche photodiode (APD) by another imaging lens (IL). The start and stop pulses are provided by the avalanche photodiode and the laser electronics, respectively (reverse mode).

Fig. 2
Fig. 2

Normalized ensemble fluorescence decay of CdSe/ZnS QDs (1.8-nm radius, 575-nm peak emission). The solid curve is a triple exponential fit to the data with components at 3.4, 16.1, and 35.6  ns. The measured fluorescence decay of Rhodamine molecules in water is also displayed (dashed curve) and is well described by a single exponential decay with time constant 4.3  ns.

Fig. 3
Fig. 3

Time-resolved confocal image of a fixed 3T3 cell: a, Image obtained from all the detected photons; b, gated image constructed from only photons that arrived 35–65  ns after the laser pulse. The image was taken with a laser intensity of 0.1 kW/cm2 and 25-ms integration time per pixel. The scale bars are 5 μm. The insets show cross sections along the same horizontal line (indicated by the black arrows) for a and b.

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