Fluorescence lifetime imaging using a single photon avalanche diode array sensor

Single photon detectors allows us work with the weakest fluorescence signals. Single photon arrays, combined with ps-controlled gating allow us to create image maps of fluorescence lifetimes, which can be used for in-vivo discrimination of tissue activity. Here we present fluorescence lifetime imaging using the ‘SwissSPAD’ sensor, a 512-by-128-pixel array of gated single photon detectors, fabricated in a standard high-voltage 0.35 μm CMOS process. We present a protocol for spatially resolved lifetime measurements where the lifetime can be retrieved for each pixel. We demonstrate the system by imaging patterns of Fluorescein and Rhodamine B on test slides, as well as measuring mixed samples to retrieve both components of the decay lifetime. The single photon sensitivity of the sensor creates a valuable instrument to perform live cell or live animal (in vivo) measurements of the weak autofluorescent signals, for example distinguishing unlabelled free and bound NADH. Our ultimate goal is to create a real time fluorescence lifetime imaging system, possibly integrated into augmented reality goggles, which could allow immediate discrimination of in vivo tissues.