Background: Infection of mothers with Rubella virus during pregnancy can be serious; if the mother is infected within the first 20 weeks of pregnancy she is likely to have miscarriage, stillbirth, or baby with congenital rubella syndrome. This study was carried out to define Rubella virus seroprevalence in pregnancy in Osogbo, Nigeria. Methods: This study is a cross-sectional sero-survey of rubella IgG antibody among pregnant women attending antenatal clinic of Ladoke Akintola University of Technology Teaching Hospital, Osogbo, Nigeria. Socio-demographic information on participants was collected by interviewer-administered questionnaire while venous samples were collected, stored at −20°C and serum samples were screened for detection of rubella IgG antibodies using the enzyme linked immunosorbent assay. Results: Of the 200 sample evaluated for rubella Immunoglobulin G antibody, 175 (87.5%) were positive and 25 (12.5%) were negative. The result indicated prevalence of 85.7% in 15-19 year age group, 86.8% in 20-24 year age group, 89.6% in 25-29 year group, and 100% in greater than 40 year age group. Rubella IgG seroprevalence was not associated with age, gestational age, gravidity, vaccination, occupation and education. Conclusions: As the immunity gap in the studied population was high, rubella vaccination should be provided for all women of child-bearing age and children