AbstractWest Nile Virus (WNV) is a mosquito‐transmitted flavivirus, widely distributed throughout Africa, Asia and the Middle East. WNV may cause epidemics of human meningoencephalitis. The unexpected emergence of WNV (New York, 1999) and its rapid spread throughout North America during the following years caused a number of blood transfusion‐ and organ transplant‐associated transmissions of WNV. In order to estimate the potential WNV threat for Central Europe, we analyzed the anti‐WNV prevalence and WNV‐RNA incidence among 14,437 and 9,976 blood donors from Germany. There was a high rate of initially anti‐WNV reactives (5.9%), but only a few cases (0.03%) were confirmed as anti‐WNV positive by neutralization assay. No WNV‐RNA positive blood donor was identified in this study. Whereas WNV‐RNA was frequently detected in manufacturing plasma pools from the US, none was detected in pools of European or Asian origin. Virus inactivation steps integrated into the manufacturing process of plasma derivatives were shown to be sufficient to assure the WNV safety of plasma derivatives. A well‐characterized WNV reference material was prepared, showing 340 WNV‐RNA copies per infectious dose. J. Med. Virol. 80:557–563, 2008. © 2008 Wiley‐Liss, Inc.