1084 IEEE SENSORS JOURNAL, VOL. 9, NO. 9, SEPTEMBER 2009 Single-Photon Avalanche Diode CMOS Sensor for Time-Resolved Fluorescence Measurements David Stoppa, Member, IEEE, Daniel Mosconi, Lucio Pancheri, and Lorenzo Gonzo, Member, IEEE Abstract—A single-photon avalanche diode–based pixel array for the analysis of fluorescence phenomena is presented. Each pixel integrates a single photon detector combined with an active quenching circuit and a 17-bit digital events counter. On-chip master logic provides the digital control phases required by the pixel array with a full programmability of the main timing synchronisms. The pixel exhibits an average dark count rate of 3 kcps and a dynamic range of over 120-dB in time uncorrelated operation. A complete characterization of the single photon avalanche diode characteristics is reported. Time-resolved fluorescence measurements have been demonstrated by detecting the fluorescence decay of quantum-dot samples without the aid of any optical filters for excitation laser light cutoff. Index Terms—CMOS, fluorescence lifetime, single-photon avalanche diode (SPAD), time-gating. I. INTRODUCTION I N recent years, there has been a growing interest in fast and low-cost biological testing. However, existing analysis- equipment are expensive, bulky and time-consuming, making their use exploitable for research applications only. Among the many methods used for biological testing, optical detection is the most commonly used. In particular, fluorescence lifetime imaging is an investigation tool of paramount importance in imaging of molecular processes in physics and life sciences re- search [1], allowing the mapping of many cell parameters (such as pH, ion concentrations, etc.) and the detection of pathologies or DNA sequencing [2]. A typical fluorescence lifetime experiment uses a pulsed or modulated laser pulse to excite the fluorophores and the emitted light is revealed by means of intensified CCD cameras or photomultipler tubes [3], in order to achieve the required time-resolution and light sensitivity specifications. The perfor- mance of these laboratory instruments are excellent in terms of time accuracy and spatial resolution but they are expensive and bulky, while there is an increasing demand for portable and inexpensive biosensors for environmental and biomedical diagnostics. In the last few years, there have been many efforts in realizing integrated circuits aimed at biosensing applications [4], [5], [11]–[15]. The use of a CMOS integrated optical Manuscript received October 02, 2008; revised December 17, 2008; accepted February 03, 2009. Current version published August 05, 2009. The associate editor coordinating the review of this paper and approving it for publication was Dr. Krikor Ozanyan. The authors are with Fondazione Bruno Kessler—Centro per la Ricerca Scientifica e Tecnologica (FBK-irst), I-38100, Povo (TN), Italy (e-mail: stoppa@fbk.eu; mosconi@fbk.eu; pancheri@fbk.eu; lgonzo@fbk.eu). Digital Object Identifier 10.1109/JSEN.2009.2025581 sensor based on simple photodiodes has been proposed for fluorescence decay detection [5], but the time resolution was limited by the RC time constant of the photodiode. High accu- racy time resolution can be achieved exploiting single-photon avalanche diodes (SPADs) [6], but most of the efforts in the implementation of such systems reported in the literature regard the integration of the SPAD with quenching circuits, and of a simple voltage comparator for the digital conversion of the received Geiger pulse [7]–[9]. Only a few examples of SPAD arrays with integrated readout channels have been reported so far. In [10], the authors report a linear SPAD array with an in-pixel Time-to-Amplitude Converter, used to detect the pho- tons arrival time. In [12], a 128 128 pixel array and a column Time-to-Digital Converter have been demonstrated. A 16 4 SPAD pixel array with two pixel-level gated counters have been recently proposed for fluorescence lifetime measurement [13], and also integrated with micropixellated light-emitting diodes in a very compact analysis system [14]. We have recently presented a chip capable of performing time-gated fluorescence detection [15], which integrates, at the pixel level, an actively quenched SPAD and a 17-bit digital counter. The chip, which includes a 7 2-pixel array, was fabricated in a high voltage 0.35- CMOS technology. In this paper, a complete characterization of the chip pro- posed in [15] is reported, using an FPGA-based control board and a USB interface for data acquisition. Using this system, lifetime measurements of quantum dot fluorophores without the use of optical filter for laser light cutoff have been successfully performed. The time-gated fluorescence lifetime measurement technique is briefly reviewed in Section II, while an overview of the pixel architecture is carried out in Section III. The chip architecture is presented in Section IV. Finally, a complete characterization of the system is reported in Section V. II. MEASURING TECHNIQUE The adopted measuring technique is sketched in Fig. 1. A sub-ns laser pulse illuminates the biological sample containing the fluorophores, and the low signal associated with the fluo- rescence emission is detected by the sensor. The adopted mea- suring technique is based on a time-gated detection method, where the light signal is detected by using two or more obser- vation windows (OWs). Each window has an externally pro- grammable time width and can be delayed with respect to the trigger laser pulse by a user-defined time value. The time offset between the laser pulse and the beginning of the observation 1530-437X/$26.00 © 2009 IEEE