Safety of hepatitis B vaccine prepared from HBeAg positive plasma containing large amounts of HBsAg was investigated by following the relative reduction of HBV DNA in the purification steps by means of DNA-DNA hybridization. Two steps in the purification procedure were found to efficiently eliminate HBV DNA in the source material. In total, HBV DNA was calculated to be reduced by a factor of 5×10-9. From these results, it was concluded that large scale production of highly safe hepatitis B vaccine from HBeAg positive plasma was possible.