A) The carboxymethylureido modification was attached to either the 6- or the 2-position of nicotine. B) BALB/c mice were immunized s.c. with 25 μg of either unconjugated KLH, 6CMUNic-KLH or 2CMUNic-KLH adsorbed on alum adjuvant, and administered on days 0, 14 and 21. Vaccine efficacy was evaluated 7–10 days after the last immunization. Immunized mice were challenged with 0.03 mg/kg i.v. nicotine. Data are mean±SEM, n=6 each group. Statistical symbols: *p<0.05, ***p<0.001, ****p<0.0001.