In a prospective, controlled, randomized, multicenter study the immunogenicity of a single (day 0) and two (day 0, 28) booster vaccination against diphtheria were compared in subjects who had received their last diphtheria vaccination more than 10 years ago. Both short-term and long-term immunogenicity was assessed by determining diphtheria antitoxin levels four weeks after vaccination and after one and two years. 102 subjects received the first booster vaccination, and 83 were vaccinated twice. Prior to the first vaccination 27% of the subjects had a diphtheria antitoxin level below 0.01 I.U./ml; after the first booster only 5% were unprotected. The second booster did not show a significant effect, however, in 1 of the 5 subjects who were still unprotected after the first booster the second elicited an antitoxin level of more than 0.01 I.U./ml. After one and two years 7% and 8% of the subjects had diphtheria antitoxin level below 0.01 I.U./ml. A serological effect of a second booster vaccination four weeks after the first one could not be demonstrated neither after one nor after two years.