Provided are a separator, a lithium secondary battery employing the same, and a method of preparing the separator. The separator includes: a porous substrate; and a coating layer arranged on at least one surface of the porous substrate, wherein the coating layer includes a binder and inorganic particles, the binder includes an aqueous cross-linking reactive polyacrylamide-based copolymer, and the polyacrylamide-based copolymer includes at least two cross-linking reactive groups cross-linkable with each other, a weight ratio of the binder to the inorganic particles is about 1:10 to about 1:35, and a thickness of the coating layer is greater than 0.5 μm and no greater than 4 μm. The separator has significantly improved thermal resistance characteristics and low resistance, and therefore, a lithium secondary battery with improved battery stability and lifespan characteristics at the same time may be provided.