Luminescent biosensors for forensic applications based on new ultrasensitive Silicon Photomultiplier detector

Global security threats have become a major worldwide concern and their early detection represents a major challenge to current monitoring technologies. The aim of this PhD research was thus focused on the development of a new portable analytical device suitable for implementation by scientific police routine screenings and inspections. The suitability of different portable light detectors was investigated, including charge coupled devices (CCDs), smartphone integrated Complementary Metal-Oxide-Semiconductor (CMOSs) and Silicon Photomultiplier (SiPM) technology. SiPM was selected thanks to its high sensitivity combined with very low power supply (few tens of volts). The ArduSiPM, a SiPM detector for the Arduino DUE microcontroller, was exploited to detect bioluminescence and chemiluminescence. A new portable device for low-light detection, named LuminoSiPM, was fabricated by 3D printing together with disposable sample-holders for liquid samples and origami paper-based biosensors. Several parameters were optimized to reduce dark noise and improve signal to noise ratio. A comparison of the LuminoSiPM analytical performance, in terms of delectability and wavelength sensitivity dependence, was performed with benchtop PMT instrumentation (Varioskan Flash), a cooled CCD sensor (ATIK 383L), and OnePlus 6 smartphone integrated CMOS, using two luciferases/luciferin system emitting a different wavelength: the green-emitting P. pyralis mutant PpyGRT (λmax 550 nm) and the blue-emitting NanoLuc (λmax 490 nm). A limit of detection of 5.3 x 10-9 M was obtained with Nanoluc, about one order of magnitude lower than those obtained with the ATIK 383L CCD camera. These promising results have allowed the development of portable biosensors based of this new device.As proof-of-principle, a paper-based origami chemiluminescent enzyme biosensor for forensic application has been developed. This biosensor is based on the inhibition of acetylcholinesterase (AChE) activity by molecules, such as organophosphate pesticides and nerve agents and, combined with LuminoSiPM, showed useful for bio-chemiluminescence detections at the point-of-need in forensic investigations.