A new method, a 3D printing technique, in particular, selective laser melting (SLM), has been used to fabricate moulds for the injection moulding of thermoplastic microfluidic chips that are suitable for prototyping and early stage scale-up. The micro metallic patterns are printed on to a pre-finished substrate to form a microstructured mould. The dimensional accuracy, surface morphology, bonding strength between the printed patterns and substrate, as well as the microstructure of micro features were all characterized. A microfluidic mould was successfully printed and used directly for injection moulding of cyclic olefin copolymer (COC) microfluidic chips, which were used subsequently to successfully monitor nitrite concentrations in environmental water. The characterization indicated that this new process can be used for fast fabrication of mould tools for injection moulding/hot embossing microfluidic devices. It is faster, more flexible and less expensive than conventional micro-machining processes, although the accuracy and finish are still needed to improve though process optimization and hybrid SLM and machining processes.