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125 results on '"Labanca, Ivan"'

1. Optical crosstalk in SPAD arrays for high-throughput single-molecule fluorescence spectroscopy

Author

Ingargiola, Antonino, Segal, Maya, Gulinatti, Angelo, Rech, Ivan, Labanca, Ivan, Maccagnani, Piera, Ghioni, Massimo, Weiss, Shimon, and Michalet, Xavier

Subjects
Nuclear and Plasma Physics, Physical Sciences, Biotechnology, SPAD array, Optical crosstalk, Single-molecule, Fluorescence, Spectroscopy, Astronomical and Space Sciences, Atomic, Molecular, Nuclear, Particle and Plasma Physics, Other Physical Sciences, Nuclear & Particles Physics, Nuclear and plasma physics
Abstract

Single-molecule fluorescence spectroscopy (SMFS), based on the detection of individual molecules freely diffusing through the excitation spot of a confocal microscope, has allowed unprecedented insights into biological processes at the molecular level, but suffers from limited throughput. We have recently introduced a multispot version of SMFS, which allows achieving high-throughput SMFS by virtue of parallelization, and relies on custom silicon single-photon avalanche diode (SPAD) detector arrays. Here, we examine the premise of this parallelization approach, which is that data acquired from different spots is uncorrelated. In particular, we measure the optical crosstalk characteristics of the two 48-pixel SPAD arrays used in our recent SMFS studies, and demonstrate that it is negligible (crosstalk probability ≤ 1.1 10-3) and undetectable in cross-correlation analysis of actual single-molecule fluorescence data.

Published
2018

2. 48-spot single-molecule FRET setup with periodic acceptor excitation

Author

Ingargiola, Antonino, Segal, Maya, Gulinatti, Angelo, Rech, Ivan, Labanca, Ivan, Maccagnani, Piera, Ghioni, Massimo, Weiss, Shimon, and Michalet, Xavier

Subjects
Physical Sciences, Chemical Sciences, Physical Chemistry, Genetics, DNA, Fluorescence Resonance Energy Transfer, Fluorescent Dyes, Kinetics, Time Factors, Engineering, Chemical Physics, Chemical sciences, Physical sciences
Abstract

Single-molecule Förster resonance energy transfer (smFRET) allows measuring distances between donor and acceptor fluorophores on the 3-10 nm range. Solution-based smFRET allows measurement of binding-unbinding events or conformational changes of dye-labeled biomolecules without ensemble averaging and free from surface perturbations. When employing dual (or multi) laser excitation, smFRET allows resolving the number of fluorescent labels on each molecule, greatly enhancing the ability to study heterogeneous samples. A major drawback to solution-based smFRET is the low throughput, which renders repetitive measurements expensive and hinders the ability to study kinetic phenomena in real-time. Here we demonstrate a high-throughput smFRET system that multiplexes acquisition by using 48 excitation spots and two 48-pixel single-photon avalanche diode array detectors. The system employs two excitation lasers allowing separation of species with one or two active fluorophores. The performance of the system is demonstrated on a set of doubly labeled double-stranded DNA oligonucleotides with different distances between donor and acceptor dyes along the DNA duplex. We show that the acquisition time for accurate subpopulation identification is reduced from several minutes to seconds, opening the way to high-throughput screening applications and real-time kinetics studies of enzymatic reactions such as DNA transcription by bacterial RNA polymerase.

Published
2018

4. Optical crosstalk in SPAD arrays for high-throughput single-molecule fluorescence spectroscopy

Author

Ingargiola, Antonino, Segal, Maya, Gulinatti, Angelo, Rech, Ivan, Labanca, Ivan, Maccagnani, Piera, Ghioni, Massimo, Weiss, Shimon, and Michalet, Xavier

Abstract

Single-molecule fluorescence spectroscopy (SMFS), based on the detection of individual molecules freely diffusing through the excitation spot of a confocal microscope, has allowed unprecedented insights into biological processes at the molecular level, but suffers from limited throughput. We have recently introduced a multispot version of SMFS, which allows achieving high-throughput SMFS by virtue of parallelization, and relies on custom silicon single-photon avalanche diode (SPAD) detector arrays. Here, we examine the premise of this parallelization approach, which is that data acquired from different spots is uncorrelated. In particular, we measure the optical crosstalk characteristics of the two 48-SPAD arrays used in our recent SMFS studies, and demonstrate that it is negligible (crosstalk probability ≤ 1.1 10 −3 ) and is undetectable in cross-correlation analysis of actual single-molecule fluorescence data.

Published
2017

5. 48-spot single-molecule FRET setup with periodic acceptor excitation

Author

Ingargiola, Antonino, Segal, Maya, Gulinatti, Angelo, Rech, Ivan, Labanca, Ivan, Maccagnani, Piera, Ghioni, Massimo, Weiss, Shimon, and Michalet, Xavier

Subjects
Genetics
Abstract

Single-molecule FRET (smFRET) allows measuring distances between donor and acceptor fluorophores on the 3-10 nm range. Solution-based smFRET allows measurement of binding-unbinding events or conformational changes of dye-labeled biomolecules without ensemble averaging and free from surface perturbations. When employing dual (or multi) laser exci-tation, smFRET allows resolving the number of fluorescent labels on each molecule, greatly enhancing the ability to study heterogeneous samples. A major drawback to solution-based smFRET is the low throughput, which renders repetitive measurements expensive and hin-ders the ability to study kinetic phenomena in real-time. Here we demonstrate a high-throughput smFRET system which multiplexes acquisition by using 48 excitation spots and two 48-pixel SPAD array detectors. The system employs two excitation lasers allowing separation of species with one or two active fluorophores. The performance of the system is demonstrated on a set of doubly-labeled double-stranded DNA oligonucleotides with different distances between donor and acceptor dyes along the DNA duplex. We show that the acquisition time for accurate subpopulation identification is reduced from several minutes to seconds, opening the way to high-throughput screening applications and real-time kinetics studies of enzymatic reactions such as DNA transcription by bacterial RNA polymerase.

Published
2017

9. A 4.5 Ps Precision TCSPC System: Design Principles and Characterization

Author

Farina, Serena, Labanca, Ivan, Acconcia, Giulia, and Rech, Ivan

Abstract

With the recent advancements in single-photon detectors, very low-jitter timing systems are required to fully exploit their performance in real applications. In this article, we present the design principles and experimental characterization of a single-channel time-correlated single-photon counting (TCSPC) system, that achieves a jitter down to 4.5 ps FWHM, a peak-to-peak differential nonlinearity of 1.5% LSB and a count rate of 12 Mcps over a nanoseconds full-scale range. These results have been attained by minimizing the different jitter contributions that are introduced at various levels in the whole timing chain, still without trading them off with the other performance parameters. To the best of our knowledge, this work represents the state-of-the-art performance in case of a full-scale range as large as 12.5 ns.

Published
2024
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28. A 150nm fully integrated active quenching circuit driving custom technology SPAD at 250Mcps

Author

Buller, Gerald S., Hollins, Richard C., Lamb, Robert A., Laurenzis, Martin, Salmon, Neil A., Gumbmann, Frank, Zamboni, Roberto, Andraud, Chantal, Szep, Attila A., Alsing, Paul M., Fanto, Michael L., Rarity, John G., Camposeo, Andrea, Persano, Luana, Busse, Lynda E., Farsari, Maria, Giudici, Andrea, Acconcia, Giulia, Labanca, Ivan, Ghioni, Massimo, and Rech, Ivan

Published
2022
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38. Above pile-up fluorescence microscopy with a 32 Mc/s single-channel time-resolved SPAD system

Author

Farina, Serena, Labanca, Ivan, Acconcia, Giulia, Ghezzi, Alberto, Farina, Andrea, D’Andrea, Cosimo, and Rech, Ivan

Abstract

One of the major drawbacks of time-correlated single-photon counting (TCSPC) is generally represented by pile-up distortion, which strongly bounds the maximum acquisition speed to a few percent of the laser excitation rate. Based on a previous theoretical analysis, recently we presented the first, to the best of our knowledge, low-distortion and high-speed TCSPC system capable of overcoming the pile-up limitation by perfectly matching the single-photon avalanche diode (SPAD) dead time to the laser period. In this work, we validate the proposed system in a standard fluorescence measurement by comparing experimental data with the reference theoretical framework. As a result, a count rate of 32 Mc/s was achieved with a single-channel system still observing a negligible lifetime distortion.

Published
2022

48. Underwater time of flight depth imaging using an asynchronous linear single photon avalanche diode detector array

Author

Buller, Gerald S., Hollins, Richard C., Lamb, Robert A., Laurenzis, Martin, Camposeo, Andrea, Farsari, Maria, Persano, Luana, Busse, Lynda E., Alsing, Paul M., Dušek, Miloslav, Fanto, Michael L., Rarity, John G., Kajzar, Francois, Szep, Attila, Zamboni, Roberto, Maccarone, Aurora, Acconcia, Giulia, Steinlehner, Ulrich, Labanca, Ivan, Rech, Ivan, and Buller, Gerald S.

Published
2020
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