Detonation propagation in a stratified layer of combustible gas over an inert gas was investigated experimentally. The layer formed in a 12.7-mm-wide channel by opening a sliding door that initially separated a nitrogen-diluted stoichiometric hydrogen–oxygen mixture from argon, or nitrogen. As the lighter combustible gas layer spreads axially down the channel, diffusion across the interface produces a composition gradient across the layer height. A steady detonation wave, generated by deflagration-to-detonation transition in the driver section before the door location, was transmitted into the combustible layer. The axial distance the layer spreads and the amount of mass diffusion across the layer were controlled by the flame ignition delay time after the door opens. Schlieren video and soot foils were used to measure the extent of detonation propagation through the layer. It was shown that detonation propagation through the layer is self-limiting due to over-mixing at the layer leading edge. Three-dimensional numerical simulations, including viscous and multicomponent mass diffusion effects, predicted the composition distribution within the layer. The cell size distribution, calculated based on the theoretical ZND induction zone length, corresponding to the simulation composition distribution showed that a cell size gradient-based failure criterion successfully predicted the extent of propagation in the layer.