Prion diseases are characterized by the accumulation of a pathological form of prion protein (PrPSc), which behaves as an infectious agent. Here we developed an in vitro co-culture system to analyze the PrPSc transmission from ScN2a cell, which persistently retains PrPSc, to naive N2a cell. In this cell-to-cell transmission system, PrPSc transmitted to recipient N2a cell was able to be detected within 5-7 days. Further characterization showed that higher cell density greatly facilitated the transmission of PrPSc. This improved in vitro transmission method may become a useful tool for unveiling the molecular mechanism of PrPSc transmission. (C) 2010 Elsevier Inc. All rights reserved.