Bioluminescence has been widely used for important biosensing applications such as the measurement of adenosine triphosphate (ATP), the energy unit in biological systems and an indicator of vital processes. The current technology for detection is mainly based on large equipment such as readers and imaging systems, which require intensive and time-consuming procedures. A miniaturised bioluminescence sensing system, which would allow sensitive and continuous monitoring of ATP, with an integrated and low-cost disposable microfluidic chamber for handling of biological samples, is highly desirable. Here, we report the design, fabrication and testing of 3D printed microfluidics chips coupled with silicon photomultipliers (SiPMs) for high sensitive real-time ATP detection. The 3D microfluidic chip reduces reactant consumption and facilitates solution delivery close to the SiPM to increase the detection …