We investigated the formation behavior of supported lipid bilayers (SLBs) on polyelectrolyte multilayers (PEMs) prepared using the layer-by-layer method. The SLBs were formed using the liposome fusion method, which was driven by electrostatic interactions. We used three types of cationic polyelectrolytes to prepare PEMs: poly(diallyldimethylammonium chloride), which consists of a linear short polymer chain with quaternary ammonium cations, polyvinylamidin, which presents a linear, long polymer chain and polyethyleneimine which features a branched polymer chain. Poly(sodium 4-styrenesulfonate) was used as an anionic polyelectrolyte. First, we evaluated the effect of the molecular structure of the polyelectrolytes on the formation of SLBs. The formation of SLBs was evaluated using water permeability data, the lateral diffusivity of lipid molecules on the PEMs was determined using a fluorescence recovery after photo-bleaching assay and the amount of lipid molecules adsorbed on the PEMs. We revealed that both the molecular structure and charge density of the polyelectrolytes affected the formation of SLBs. Furthermore, we could form SLBs on high permeable PEMs by combining different cationic polyelectrolytes. These SLBs would be applicable for a platform to immobilize lipophilic biomolecules. (C) 2020 Elsevier Inc. All rights reserved.