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Abstract: In the field of genomic research, the microcapillary electrophoresis (MCE) plays a key role, allowing a small reagent use but requiring the ability to detect luminescences down to a few photons per second. Very high sensitivity detectors are therefore required, rising the systems’ final costs and reducing their diffusion. There is a strong request to develop compact and low cost systems. The state of the art for MCE-based DNA analysis instruments is the use of photomultiplier tubes (PMTs) or CCDs, but either have some drawbacks. As for the PMTs, they cannot be integrated, so detection of the four bases can occur only using four different PMTs, increasing size and cost. As for the CCDs, even if they are commonly used, they are not the best solution regarding simplicity and cost. A valid alternative is single photon avalanche diodes (SPADs). Such devices present high quantum efficiency (higher than PMTs), with the possibility to integrate several detectors on a single chip, even with custom geometries, retaining low fabrication costs and high sensitivity. The strong potential of planar SPAD technology for DNA analysis has already been demonstrated in previous works. Here we present the first genotyping system based on a custom array of four SPADs, purposely designed to be used in conjunction with Beckman-Coulter markers. The module size is 19cm (L), 12.5cm (W), 15cm (H), including high voltage section, detectors and optical system: at the best of our knowledge, this is the most compact four-colour genotyping system in literature. [Copyright &y& Elsevier]

Inductor current sensing is becoming widely used in current programmed controllers for microprocessor applications. This method exploits a low-pass filter in parallel with the inductor to provide lossless current sense. A major drawback of inductor. current sensing is that accurate sense the dc and ac components of the current signal requires precise matching between the low-pass filter time constant and the inductor time constant (L/R[subL]). However, matching accuracy depends on the tolerance of the components and on the operating conditions; therefore it can hardly be guaranteed. To overcome this problem, a novel digital autotuning system is proposed that automatically compensates any time constant mismatch. This autotuning system has been developed for voltage regulator module (VRM) current programmed controllers. It makes it possible to meet the adaptive voltage positioning requirements using conventional and low cost components, and to solve problems such as aging effects, temperature variations, and process tolerances as well. A prototype of the autotuning system based on a field-programmable gate array and a commercial dc/dc controller has been designed and tested. The experimental results fully confirmed the effectiveness of the proposed method, showing an improvement of the current sense precision from about 30% up to 4%. This innovative solution is suitable to fulfill the challenging accuracy specifications required by the future VRM applications. [ABSTRACT FROM AUTHOR]

This paper investigates a mixed synchronous/asynchronous digital voltage-mode controller for DC-DC converters. In the proposed control architecture, the turn-on switching event is determined asynchronously by comparing the converter output voltage and a synchronously generated voltage ramp driven by the digital control using a low-resolution digital-to-analog converter. Switch turn-off is determined synchronously by the system clock. In the proposed approach, the derivative action of the proportional-integral-derivative voltage-mode controller is inherently obtained by the frequency modulation, without requiring the digital computation of the derivative action. A simplified small-signal model is also derived in order to analyze the performance achievable by the proposed solution. This control architecture features good dynamic performance, and frequency modulation during transients. Simulation and experimental results on a synchronous buck converter, where the digital control has been implemented in field programmable gate array, confirm the effectiveness of the proposed solution. [ABSTRACT FROM AUTHOR]

This paper proposes a simple autotuning technique for digitally controlled dc-dc converters. The proposed approach is based on the relay feedback method and introduces perturbations on the output voltage during converter soft-start. By using an iterative procedure, the tuning of proportional-integral-derivative parameters is obtained directly by including the controller in the relay feedback and by adjusting the controller parameters based on the specified phase margin and control loop bandwidth. A nice property of the proposed solution is that output voltage perturbations are introduced while maintaining the relay feedback control on the output voltage. The proposed algorithm is simple, requires small tuning times, and it is compliant with the cost/complexity constraints of integrated digital integrated circuits. Simulation and experimental results of a synchronous buck converter and of a dc-dc boost converter confirm the effectiveness of the proposed solution. [ABSTRACT FROM AUTHOR]

Design and fabrication of single photon avalanche detector (SPAD) in planar technology is reported. Device design and critical issues in the technology are discussed. Experimental test procedures are described for dark-counting rate, afterpulsing probability, photon timing resolution, and quantum detection efficiency. Low-noise detectors are obtained, with dark counting rates down to 10 c/s for devices with 10 µm diameter, down to 1 kc/s for 50 µm diameter. The technology is suitable for monolithic integration of SPAD detectors and associated circuits. [ABSTRACT FROM AUTHOR]

Timing measurements with silicon photomultipliers (SiPMs) are affected by the major issue of the so-called time-walk effect; this is due to the intrinsic architecture of SiPM devices. A new technique for time-walk compensation in timing measurements with SiPMs is presented and discussed, feasible with a low-complexity and low-cost circuitry and able to improve the timing accuracy. The validity of the presented technique has been experimentally proven with a 3 × 3 mm2 Hamamatsu MPPC, achieving a jitter reduction of a factor 2.85 compared with the case without time-walk compensation. To perform the experimental validation, a discrete-components transimpedance amplifier is developed with large bandwidth and low jitter, specifically designed to read-out and amplify the SiPM signal. [ABSTRACT FROM AUTHOR]

At the present time, Single Photon Avalanche Diodes (SPADs) are the enabling devices in many applications, ranging from medical imaging to laser ranging and from remote sensing to quantum key distribution. Even though they belong to different scientific domains, these applications share the need for a detector capable of attaining high count rates possibly without trading it off with other key detector's features, such as afterpulsing probability, photon detection efficiency, and dark counts. In this work, we present the characterization of a fast integrated active quenching circuit capable of driving high-performance external custom-technology SPADs for single photon detection in the visible wavelength range. Combining the prompt intervention of the electronic circuitry and the performance of a custom-technology SPAD, we attained count rates up to 250 MCps while keeping the afterpulsing probability within 2%.

Single-photon detection is an invaluable tool for many applications ranging from basic research to consumer electronics. In this respect, the Single Photon Avalanche Diode (SPAD) plays a key role in enabling a broad diffusion of these techniques thanks to its remarkable performance, room-temperature operation, and scalability. In this paper we present a silicon technology that allows the fabrication of SPAD-arrays with an unprecedented combination of low timing jitter (95 ps FWHM) and high detection efficiency at red and near infrared wavelengths (peak of 70% at 650 nm, 45% at 800 nm). We discuss the device structure, the fabrication process, and we present a thorough experimental characterization of the fabricated detectors. We think that these results can pave the way to new exciting developments in many fields, ranging from quantum optics to single molecule spectroscopy.

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.

Time-Correlated Single Photon Counting (TCSPC) is an essential tool in many scientific applications, where the recording of optical pulses with picosecond precision is required. Unfortunately, a key issue has to be faced: distortion phenomena can affect TCSPC experiments at high count rates. In order to avoid this problem, TCSPC experiments have been commonly carried out by limiting the maximum operating frequency of a measurement channel below 5% of the excitation frequency, leading to a long acquisition time. Recently, it has been demonstrated that matching the detector dead time to the excitation period allows to keep distortion around zero regardless of the rate of impinging photons. This solution paves the way to unprecedented measurement speed in TCSPC experiments. In this scenario, the front-end circuits that drive the detector play a crucial role in determining the performance of the system, both in terms of measurement speed and timing performance. Here we present two fully integrated front-end circuits for Single Photon Avalanche Diodes (SPADs): a fast Active Quenching Circuit (AQC) and a fully-differential current pick-up circuit. The AQC can apply very fast voltage variations, as short as 1.6ns, to reset external custom-technology SPAD detectors. A fast reset, indeed, is a key parameter to maximize the measurement speed. The current pick-up circuit is based on a fully differential structure which allows unprecedented rejection of disturbances that typically affect SPAD-based systems at the end of the dead time. The circuit permits to sense the current edge resulting from a photon detection with picosecond accuracy and precision even a few picoseconds after the end of the dead time imposed by the AQC. This is a crucial requirement when the system is operated at high rates. Both circuits have been deeply characterized, especially in terms of achievable measurement speed and timing performance.

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.

In this letter, the development and the experimental characterization of a new photon detection module, based on a 32×1 red-enhanced single-photon avalanche diode (RE-SPAD) array, are presented. A custom-developed technology has been exploited to design a detector having large-area pixels (50-µm diameter) with optimized performance. With an excess bias voltage V oυ = 15 V, a photon detection efficiency as high as 57% at 600 nm (33% at 800 nm) is achieved, along with dark count rate in the kHz range and optical crosstalk probability as low as 0.29%. The remarkable detection efficiency of the RE-SPAD array makes the module particularly suitable for all applications where high detection efficiency in the red/near-infrared range is mandatory. As an example, the performance of the array module is demonstrated to match the demanding requirements of multispot single-molecule fluorescence spectroscopy., Competing Interests: Conflict of interest statement: M. Ghioni discloses equity in Micro Photon Devices S.r.l. (MPD).

In recent years the development of Single-Photon Avalanche Diodes (SPADs) had a big impact on single-photon counting applications requiring high-performance detectors in terms of Dark Count Rate (DCR), Photon Detection Efficiency (PDE), afterpulsing probability, etc. Among these, it is possible to find applications in single-molecule fluorescence spectroscopy that suffer from long-time measurements. In these cases SPAD arrays can be a solution in order to shorten the measurement time, thanks to the high grade of parallelism they can provide. Moreover, applications in other fields (e.g. astronomy) demand for large-area single-photon detectors, able also to handle very high count rates. For these reasons we developed a new single-photon detection module, featuring an 8 × 8 SPAD array. Thanks to a dedicated silicon technology, the performance of the detector have been finely optimized, reaching a 49% detection efficiency at 550 nm, as well as low dark counts (2 kcount/s maximum all over the array). This module can be used in two different modes: the first is a multi-spot configuration, allowing the acquisition of 64 optical signals at the same time and considerably reducing the time needed for a measurement. The second operation mode instead exploits all the pixels in a combined mode, allowing the detection of a 64-times higher maximum photon rate (up to 2 Gcount/s). In addition, this configuration provides also an extended dynamic range and allows to attain photon number resolving capabilities. Dark counts, detection efficiency, linearity, afterpulsing and crosstalk probability have been characterized at different operating conditions., Competing Interests: Conflict of interest statement: M. Ghioni discloses equity in Micro Photon Devices S.r.l. (MPD).

We describe an 8-spot confocal setup for high-throughput smFRET assays and illustrate its performance with two characteristic experiments. First, measurements on a series of freely diffusing doubly-labeled dsDNA samples allow us to demonstrate that data acquired in multiple spots in parallel can be properly corrected and result in measured sample characteristics consistent with those obtained with a standard single-spot setup. We then take advantage of the higher throughput provided by parallel acquisition to address an outstanding question about the kinetics of the initial steps of bacterial RNA transcription. Our real-time kinetic analysis of promoter escape by bacterial RNA polymerase confirms results obtained by a more indirect route, shedding additional light on the initial steps of transcription. Finally, we discuss the advantages of our multispot setup, while pointing potential limitations of the current single laser excitation design, as well as analysis challenges and their solutions.

Single-molecule spectroscopy on freely-diffusing molecules allows detecting conformational changes of biomolecules without perturbation from surface immobilization. Resolving fluorescence lifetimes increases the sensitivity in detecting conformational changes and overcomes artifacts common in intensity-based measurements. Common to all freely-diffusing techniques, however, are the long acquisition times. We report a time-resolved multispot system employing a 16-channel SPAD array and TCSPC electronics, which overcomes the throughput issue. Excitation is obtained by shaping a 532 nm pulsed laser into a line, matching the linear SPAD array geometry. We show that the line-excitation is a robust and cost-effective approach to implement multispot systems based on linear detector arrays.

Single photon avalanche diodes (SPADs) have been subject to a fast improvement in recent years. In particular, custom technologies specifically developed to fabricate SPAD devices give the designer the freedom to pursue the best detector performance required by applications. A significant breakthrough in this field is represented by the recent introduction of a red enhanced SPAD (RE-SPAD) technology, capable of attaining a good photon detection efficiency in the near infrared range (e.g. 40% at a wavelength of 800 nm) while maintaining a remarkable timing resolution of about 100ps full width at half maximum. Being planar, the RE-SPAD custom technology opened the way to the development of SPAD arrays particularly suited for demanding applications in the field of life sciences. However, to achieve such excellent performance custom SPAD detectors must be operated with an external active quenching circuit (AQC) designed on purpose. Next steps toward the development of compact and practical multichannel systems will require a new generation of monolithically integrated AQC arrays. In this paper we present a new, fully integrated AQC fabricated in a high-voltage 0.18 µm CMOS technology able to provide quenching pulses up to 50 Volts with fast leading and trailing edges. Although specifically designed for optimal operation of RE-SPAD devices, the new AQC is quite versatile: it can be used with any SPAD detector, regardless its fabrication technology, reaching remarkable count rates up to 80 Mcounts/s and generating a photon detection pulse with a timing jitter as low as 119 ps full width at half maximum. The compact design of our circuit has been specifically laid out to make this IC a suitable building block for monolithically integrated AQC arrays.

In this letter we present a compact photon detection module, based on an 8×8 array of single-photon avalanche diodes (SPADs). The use of a dedicated silicon technology for the fabrication of the sensors allows us to combine large active areas (50-μm diameter), high photon detection efficiency (49% at 550-nm wavelength) and low dark count rate. Thanks to a fully parallel architecture, the module provides voltage pulses synchronous to each photon detection for a maximum global count rate exceeding 1 Gcps. These properties makes the system suitable for operation in two different free-running modes. The first, suitable to acquire faint signals, allows multi-spot acquisitions and can be used to considerably reduce the measurement time in applications like single-molecule analysis. With the second it is possible to use all the pixels in a combined mode, to extend and move the dynamic range of the module to very high count rates and to attain number resolving capabilities.

In order to fulfill the requirements of many applications, we recently developed a new technology aimed at combining the advantages of traditional thin and thick silicon Single Photon Avalanche Diodes (SPAD). In particular we demonstrated single-pixel detectors with a remarkable improvement in the Photon Detection Efficiency in the red/near-infrared spectrum (e.g. 40% at 800nm) while maintaining a timing jitter better than 100ps. In this paper we discuss the limitations of such Red-Enhanced (RE) technology from the point of view of the fabrication of small arrays of SPAD and we propose modifications to the structure aimed at overcoming these issues. We also report the first preliminary experimental results attained on devices fabricated adopting the improved structure., Competing Interests: M. Ghioni discloses equity in Micro Photon Devices S.r.l. (MPD).

Solution-based single-molecule fluorescence spectroscopy is a powerful experimental tool with applications in cell biology, biochemistry and biophysics. The basic feature of this technique is to excite and collect light from a very small volume and work in a low concentration regime resulting in rare burst-like events corresponding to the transit of a single molecule. Detecting photon bursts is a challenging task: the small number of emitted photons in each burst calls for high detector sensitivity. Bursts are very brief, requiring detectors with fast response time and capable of sustaining high count rates. Finally, many bursts need to be accumulated to achieve proper statistical accuracy, resulting in long measurement time unless parallelization strategies are implemented to speed up data acquisition. In this paper we will show that silicon single-photon avalanche diodes (SPADs) best meet the needs of single-molecule detection. We will review the key SPAD parameters and highlight the issues to be addressed in their design, fabrication and operation. After surveying the state-of-the-art SPAD technologies, we will describe our recent progress towards increasing the throughput of single-molecule fluorescence spectroscopy in solution using parallel arrays of SPADs. The potential of this approach is illustrated with single-molecule Förster resonance energy transfer measurements.

In order to fulfill the requirements of many applications, we recently developed a new technology aimed at combining the advantages of traditional thin and thick silicon Single Photon Avalanche Diodes (SPAD). In particular we demonstrated single-pixel detectors with a remarkable improvement in the Photon Detection Efficiency at the longer wavelengths (e.g. 40% at 800nm) while maintaining a timing jitter better than 100ps. In this paper we will analyze the factors the currently prevent the fabrication of arrays of SPADs by adopting such a Red-Enhanced (RE) technology and we will propose further modifications to the device structure that will enable the fabrication of high performance RE-SPAD arrays for photon timing applications.

Single-molecule Förster resonance energy transfer (smFRET) techniques are now widely used to address outstanding problems in biology and biophysics. In order to study freely diffusing molecules, current approaches consist in exciting a low concentration (<100 pM) sample with a single confocal spot using one or more lasers and detecting the induced single-molecule fluorescence in one or more spectrally- and/or polarization-distinct channels using single-pixel Single-Photon Avalanche Diodes (SPADs). A large enough number of single-molecule bursts must be accumulated in order to compute FRET efficiencies with sufficient statistics. As a result, the minimum timescale of observable phenomena is set by the minimum acquisition time needed for accurate measurements, typically a few minutes or more, limiting this approach mostly to equilibrium studies. Increasing smFRET analysis throughput would allow studying dynamics with shorter timescales. We recently demonstrated a new multi-spot excitation approach, employing a novel multi-pixel SPAD array, using a simplified dual-view setup in which a single 8-pixel SPAD array was used to collect FRET data from 4 independent spots. In this work we extend our results to 8 spots and use two 8-SPAD arrays to collect donor and acceptor photons and demonstrate the capabilities of this system by studying a series of doubly labeled dsDNA samples with different donor-acceptor distances ranging from low to high FRET efficiencies. Our results show that it is possible to enhance the throughput of smFRET measurements in solution by almost one order of magnitude, opening the way for studies of single-molecule dynamics with fast timescale once larger SPAD arrays become available.

In this paper we present an array of 48 Single Photon Avalanche Diodes (SPADs) specifically designed for multispot Single Molecule Analysis. The detectors have been arranged in a 12×4 square geometry with a pitch-to-diameter ratio of ten in order to minimize the collection of the light from non-conjugated excitation spots. In order to explore the trade-offs between the detectors' performance and the optical coupling with the experimental setup, SPADs with an active diameter of 25 and of 50µm have been manufactured. The use of a custom technology, specifically designed for the fabrication of the detectors, allowed us to combine a high photon detection efficiency (peak close to 50% at a wavelength of 550nm) with a low dark count rate compatible with true single molecule detection. In order to allow easy integration into the optical setup for parallel single-molecule analysis, the SPAD array has been incorporated in a compact module containing all the electronics needed for a proper operation of the detectors.

Single-molecule fluorescence spectroscopy of freely diffusing molecules in solution is a powerful tool used to investigate the properties of individual molecules. Single-Photon Avalanche Diodes (SPADs) are the detectors of choice for these applications. Recently a new type of SPAD detector was introduced, dubbed red-enhanced SPAD (RE-SPAD), with good sensitivity throughout the visible spectrum and with excellent timing performance. We report a characterization of this new detector for single-molecule fluorescence resonant energy transfer (smFRET) studies on freely diffusing molecules in a confocal geometry and alternating laser excitation (ALEX) scheme. We use a series of doubly-labeled DNA molecules with donor-to-acceptor distances covering the whole range of useful FRET values. Both intensity-based (μs-ALEX) and lifetime-based (ns-ALEX) measurements are presented and compared to identical measurements performed with standard thick SPADs. Our results demonstrate the great potential of this new detector for smFRET measurements and beyond.

Light is strictly connected with life, and its presence is fundamental for any living environment. Thus, many biological mechanisms are related to light interaction or can be evaluated through processes involving energy exchange with photons. Optics has always been a precious tool to evaluate molecular and cellular mechanisms, but the discovery of lasers opened new pathways of interactions of light with biological matter, pushing an impressive development for both therapeutic and diagnostic applications in biomedicine. The use of light in different fields has become so widespread that the word photonics has been utilized to identify all the applications related to processes where the light is involved. The photonics area covers a wide range of wavelengths spanning from soft X-rays to mid-infrared and includes all devices related to photons as light sources, optical fibers and light guides, detectors, and all the related electronic equipment. The recent use of photons in the field of telecommunications has pushed the technology toward low-cost, compact, and efficient devices, making them available for many other applications, including those related to biology and medicine where these requirements are of particular relevance. Moreover, basic sciences such as physics, chemistry, mathematics, and electronics have recognized the interdisciplinary need of biomedical science and are translating the most advanced researches into these fields. The Politecnico school has pioneered many of them,and this article reviews the state of the art of biomedical research at the Politecnico in the field internationally known as biophotonics.

We present a novel approach to high-throughput Fluorescence Correlation Spectroscopy (FCS) which enables us to obtain one order of magnitude improvement in acquisition time. Our approach utilizes a liquid crystal on silicon spatial light modulator to generate dynamically adjustable focal spots, and uses an eight-pixel monolithic single-photon avalanche photodiode array. We demonstrate the capabilities of this system by showing FCS of Rhodamine 6G under various viscosities, and by showing that, with proper calibration of each detection channel, one order of magnitude improvement in acquisition speed is obtained. More generally, our approach will allow higher throughput single-molecule studies to be performed.

Solution-based single-molecule spectroscopy and fluorescence correlation spectroscopy (FCS) are powerful techniques to access a variety of molecular properties such as size, brightness, conformation, and binding constants. However, this is limited to low concentrations, which results in long acquisition times in order to achieve good statistical accuracy. Data can be acquired more quickly by using parallelization. We present a new approach using a multispot excitation and detection geometry made possible by the combination of three powerful new technologies: (i) a liquid crystal spatial light modulator to produce multiple diffraction-limited excitation spots; (ii) a multipixel detector array matching the excitation pattern and (iii) a low-cost reconfigurable multichannel counting board. We demonstrate the capabilities of this technique by reporting FCS measurements of various calibrated samples as well as single-molecule burst measurements.