Miniature microscopy provides a transformative approach to observe objects and enable continuous monitoring with ultra-compact microscopes attached directly to specimens, facilitating parallel analysis. This innovation is particularly valuable for applications such as drug discovery using organ-on-a-chip devices, which require the assessment of numerous drug/sample pairs prior to clinical trials. Ultra-compact microscopes were previously limited to brightfield techniques, which prevented the use of powerful tools like fluorescent microscopy. In this work, we present a miniature microscope with integrated fluorescence measurement capabilities. This microscope consists of a custom chip with a 10 μm-diameter single-photon avalanche diode (SPAD) faced to a 640 × 480 InGaN/GaN 4 μm-pitch LED microdisplay. It operates in raster mode, activating individual LEDs to map specimens in 2D while measuring fluorescence light with the SPAD chip. Our results demonstrate its suitability for life sciences imaging. For example, we observed a muscle-ona-chip stained with Alexa Fluor 488 to study drug efficacy on sarcopenia. Furthermore, these microscopes exhibit superior speed compared to the previously reported ultra-compact brightfield microscopes, achieving a 240-fold increase in imaging rate by means of hardware controller integration on FPGA.
KEYWORDS: Sensors, Cameras, Microscopy, Light emitting diodes, Photodetectors, Signal to noise ratio, Time metrology, Statistical methods, Optical testing, Single photon
In modern single pixel microscopy techniques, like Nano-Illumination Microscopy, long measurement times can become a major issue, especially when imaging biological tissues with large field of view. Usually, light intensity measurements are performed with CMOS pixels, with typical integration times around tens of milliseconds. In this work, we propose to obtain a light intensity measurement indirectly by applying statistical techniques to the photon arrival times gathered with an SPAD photodetector. We will present how the different statistical measurements can be used to minimize the total acquisition time and minimize also the hardware required. In this work, with captures of 256 SPAD measurements, reducing measurement time from 50ms to 50us. The dynamic range is extended by combining multiple statistical techniques with standard intensity measurements. This paves the way to practical Nano-Illumination Microscopy and other single pixel microscopy techniques.
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