Clear plastic fluidic devices with ports for incorporating electrodes to enable electrochemiluminescence (ECL) measurements were prepared using a low-cost, desktop three-dimensional (3D) printer based on stereolithography. Electrodes consisted of 0.5 mm pencil graphite rods and 0.5 mm silver wires inserted into commercially available 1/4 in.-28 threaded fittings. A bioimaging system equipped with a CCD camera was used to measure ECL generated at electrodes and small arrays using 0.2 M phosphate buffer solutions containing tris(2,2′-bipyridyl)dichlororuthenium(II) hexahydrate ([Ru(bpy)3]2+) with 100 mM tri-n-propylamine (TPA) as the coreactant. ECL signals produced at pencil graphite working electrodes were linear with respect to [Ru(bpy)3]2+ concentration for 9-900 μM [Ru(bpy)3]2+. The detection limit was found to be 7 μM using the CCD camera with exposure time set at 10 s. Electrode-to-electrode ECL signals varied by ±7.5%. Device performance was further evaluated using pencil graphite electrodes coated with multilayer poly(diallyldimethylammonium chloride) (PDDA)/DNA films. In these experiments, ECL resulted from the reaction of [Ru(bpy)3]3+ with guanines of DNA. ECL produced at these thin-film electrodes was linear with respect to [Ru(bpy)3]2+ concentration from 180 to 800 μM. These studies provide the first demonstration of ECL measurements obtained using a 3D-printed closed-channel fluidic device platform. The affordable, high-resolution 3D printer used in these studies enables easy, fast, and adaptable prototyping of fluidic devices capable of incorporating electrodes for measuring ECL.