Background In Guinea-Bissau we conducted a randomized trial of OPV0 versus No OPV0 to test the effect of not receiving OPV0 on infant mortality and morbidity. In two subgroups of participants, 6-week-old children and 6-month-old children, we investigated the effect of OPV0 on neutralizing antibodies against poliovirus type 1 and 3. Design A subgroup of infants randomized to receive OPV0 or No OPV0 in addition to the usual childhood vaccines were visited at home at 6weeks or 6months of age, and a blood sample was collected from the child and the mother. Setting Urban Guinea-Bissau. Main outcome Geometric mean titers (GMT) of neutralizing antibodies and seropositivity (titer⩾1:8) for poliovirus type 1 and 3. Results OPV0 did not affect the overall seropositivity at 6weeks or 6months of age for either polio 1 or 3. In 6-week-old infants, not receiving OPV0 was associated with significantly lower GMT for polio 1 and 3 (GMT ratio=0.52 (95% CI=0.33–0.79) for polio 1; 0.44 (0.28–0.70) for polio 3), the effect being significant in its own right in boys and in children whose mothers had low antibody levels. In contrast, in 6-month-old infants, not receiving OPV0 was associated with significantly higher GMT for polio 1 (GMT ratio=2.10 (1.32–3.35)). This was significant in its own right in boys and in children of mothers with high antibody levels. Conclusions OPV0 may contribute to early polio protection, particularly in children of mothers with low antibody levels. However, OPV0 did not contribute to overall polio immunity after subsequent doses of OPV were given, and was associated with significantly lower antibody titers in children of mothers with high antibody levels. However, it did not negatively affect the proportion of seropositive children.