Philosophical Transactions of the Royal Society B: Biological Sciences
Restricted accessReview article

Development of new photon-counting detectors for single-molecule fluorescence microscopy

X. Michalet

X. Michalet

Department of Chemistry and Biochemistry, UCLA, Los Angeles, CA 90095-1547, USA

[email protected]

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R. A. Colyer

R. A. Colyer

Department of Chemistry and Biochemistry, UCLA, Los Angeles, CA 90095-1547, USA

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G. Scalia

G. Scalia

Department of Chemistry and Biochemistry, UCLA, Los Angeles, CA 90095-1547, USA

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A. Ingargiola

A. Ingargiola

Department of Chemistry and Biochemistry, UCLA, Los Angeles, CA 90095-1547, USA

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R. Lin

R. Lin

Department of Chemistry and Biochemistry, UCLA, Los Angeles, CA 90095-1547, USA

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J. E. Millaud

J. E. Millaud

Department of Chemistry and Biochemistry, UCLA, Los Angeles, CA 90095-1547, USA

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S. Weiss

S. Weiss

Department of Chemistry and Biochemistry, UCLA, Los Angeles, CA 90095-1547, USA

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Oswald H. W. Siegmund

Oswald H. W. Siegmund

Space Sciences Laboratory, UCB, Berkeley, CA 94720, USA

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Anton S. Tremsin

Anton S. Tremsin

Space Sciences Laboratory, UCB, Berkeley, CA 94720, USA

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John V. Vallerga

John V. Vallerga

Space Sciences Laboratory, UCB, Berkeley, CA 94720, USA

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A. Cheng

A. Cheng

Department of Physics and Astronomy, UCLA, Los Angeles, CA 90095-1547, USA

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M. Levi

M. Levi

Department of Physics and Astronomy, UCLA, Los Angeles, CA 90095-1547, USA

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D. Aharoni

D. Aharoni

Department of Physics and Astronomy, UCLA, Los Angeles, CA 90095-1547, USA

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K. Arisaka

K. Arisaka

Department of Physics and Astronomy, UCLA, Los Angeles, CA 90095-1547, USA

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F. Villa

F. Villa

Dipartimento di Elettronica ed Informazione, Politecnico di Milano, Milano, Italy

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F. Guerrieri

F. Guerrieri

Dipartimento di Elettronica ed Informazione, Politecnico di Milano, Milano, Italy

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F. Panzeri

F. Panzeri

Dipartimento di Elettronica ed Informazione, Politecnico di Milano, Milano, Italy

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I. Rech

I. Rech

Dipartimento di Elettronica ed Informazione, Politecnico di Milano, Milano, Italy

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A. Gulinatti

A. Gulinatti

Dipartimento di Elettronica ed Informazione, Politecnico di Milano, Milano, Italy

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F. Zappa

F. Zappa

Dipartimento di Elettronica ed Informazione, Politecnico di Milano, Milano, Italy

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M. Ghioni

M. Ghioni

Dipartimento di Elettronica ed Informazione, Politecnico di Milano, Milano, Italy

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S. Cova

S. Cova

Dipartimento di Elettronica ed Informazione, Politecnico di Milano, Milano, Italy

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Published:https://doi.org/10.1098/rstb.2012.0035

    Two optical configurations are commonly used in single-molecule fluorescence microscopy: point-like excitation and detection to study freely diffusing molecules, and wide field illumination and detection to study surface immobilized or slowly diffusing molecules. Both approaches have common features, but also differ in significant aspects. In particular, they use different detectors, which share some requirements but also have major technical differences. Currently, two types of detectors best fulfil the needs of each approach: single-photon-counting avalanche diodes (SPADs) for point-like detection, and electron-multiplying charge-coupled devices (EMCCDs) for wide field detection. However, there is room for improvements in both cases. The first configuration suffers from low throughput owing to the analysis of data from a single location. The second, on the other hand, is limited to relatively low frame rates and loses the benefit of single-photon-counting approaches. During the past few years, new developments in point-like and wide field detectors have started addressing some of these issues. Here, we describe our recent progresses towards increasing the throughput of single-molecule fluorescence spectroscopy in solution using parallel arrays of SPADs. We also discuss our development of large area photon-counting cameras achieving subnanosecond resolution for fluorescence lifetime imaging applications at the single-molecule level.

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