One of the main issues of Single Photon Avalanche Diode arrays is optical crosstalk. Since its intensity increases with reducing the distance between devices, this phenomenon limits the density of integration within arrays. In the past optical crosstalk was ascribed essentially to the light propagating from one detector to another through direct optical paths. Accordingly, reflecting trenches between devices were proposed to prevent it, but they proved to be not completely effective. In this paper we will present experimental evidence that a significant contribution to optical crosstalk comes from light reflected internally off the bottom of the chip, thus being impossible to eliminate it completely by means of trenches. We will also propose an optical model to predict the dependence of crosstalk on the distance between devices.

Single-photon detectors play a key role in many research fields such as biology, chemistry, medicine, and space technology, and in recent years, single-photon avalanche diodes (SPADs) have become a valid alternative to photo multiplier tubes (PMTs). Moreover, scientific research has recently focused on single-photon detector arrays, pushed by a growing demand for multichannel systems. In this scenario, we developed a compact 32-channel system for time-resolved single-photon counting applications. The system is divided into two independent modules: a photon detection head including a 32 Â 1 SPAD array built in custom technology, featuring high time resolution, high photon detection efficiency (44% at 550 nm), and low dark count rate (mean value G 400 cps at À10 C) at 6-V excess bias voltage and a 32-channel acquisition system able to perform timecorrelated single-photon counting (TCSPC) measurements. The TCSPC module includes eight four-channel time-to-amplitude converter (TAC) arrays, built-in 0.35-m Si-Ge BiCMOS technology, characterized by low differential non-linearity (rms value lower than 0.15% of the time bin width) and variable full-scale range. The system response function of this TCSPC instrumentation achieves a mean time resolution of 63 ps FWHM , considering a mean count rate of 1 Mcps.

Single-molecule FRET (smFRET) allows measuring distances between donor and acceptor fluorophores on the 3-10 nm range. Solution-based smFRET allows measurement of bindingunbinding 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 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.

A modification of a standard Perkin Elmer SPCM-AQR photon detector module that remarkably improves the photon timing performance is presented here. The modification consists of an additional timing circuit board, which is inserted in the module without modifying the original circuit board. The essential feature is a pulse pickup linear network, connected to the high-voltage terminal of the photodetector, which extracts a short pulse signal with fast rise, coincident with the rise of the avalanche current. The information about the photon arrival time is obtained by sensing the onset of the rise. At low counting rates (<10^5 counts/ s) time-correlated photon counting tests show that the instrumental resolution function (IRF) thus obtained has full width at half maximum (FWHM) narrower by about 40% with respect to the original module. At higher counting rate, up to few Mcounts/s, the advantage is even more remarkable: The timing circuit practically eliminates the drawbacks that plague the original module, namely, a progressive increase of the FWHM and a progressive shift of the peak position of the IRF with increasing counting rate. The modified SPCM-AQR module is therefore suitable also for applications requiring subnanosecond time resolution at high and/or variable counting rate, such as fluorescent decay measurements, fluorescent lifetime imaging, single molecule detection and spectroscopy, and optical radar techniques

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.