An ruthenium(II) complex-containing ultrathin film, layered double hydroxide/poly(sodium 4-styrene sulfonate)/ruthenium(II) tris(bipyridine)/poly(sodium 4-styrene sulfonate) (denoted as LDH/PSS/Ru(bpy)(3)(2+)/PSS), has been fabricated via the layer-by-layer (LBL) method, which was demonstrated as an solid-state electrochemiluminescence (ECL) sensor for dihydronicotinamide adenine dinucleotide (NADH). The growth of the LBL film was monitored by the UV-vis absorption spectroscopy. XRD measurements indicate the LDH/PSS/Ru(bpy)(3)(2+)/PSS ultrathin film possesses a long range stacking order in the normal direction of the substrate, with an average repeating distance of 10.7 nm. SEM images show that the film surface displays a continuous and uniform morphology, with the root-mean square (rms) roughness of similar to 9.0 nm revealed by AFM. The ECL detection for NADH in phosphate buffer solution (pH = 7.4) was performed by using the LDH/PSS/Ru(bpy)(3)(2+)/PSS film sensor, and a linear response was obtained in the range 3.16 x 10(-6) to 3.23 x 10(-4) M with a detection limit of 0.023 mu M. Therefore, it is expected that the LBL film based on ruthenium(II) complex in this work can be potentially used as an ultrasensitive ECL sensor in environmental and biological analysis. (C) 2012 Elsevier Ltd. All rights reserved.