The fabrication of conventional PDMS on glass lab-on-a-chip (LOC) devices, using templates printed with a commercial (2299 US$) micro-stereo lithography 3D printer, is demonstrated. Printed templates replace clean room and photolithographic fabrication resources and deliver resolutions of 50 μm, and up to 10 μm in localized hindrances, whereas the templates are smooth enough to allow direct transfer and proper sealing to glass substrates. 3D printed templates accommodate multiple thicknesses, from 50 μm up to several mm within the same template, with no additional processing cost or effort. This capability is exploited to integrate silicone tubing easily, to improve micromixer performance and to produce multilevel fluidics with simple access to independent functional surfaces, which is illustrated by time-resolved glucose detection. The templates are reusable, can be fabricated in under 20 min, with an average cost of 0.48 US$, which promotes broader access to established LOC configurations with minimal fabrication requirements, relieves LOC fabrication from design skills and provides a versatile LOC development platform. © The Royal Society of Chemistry.