Background: Menstruation has more profound effect on quality of education than other aspect of puberty because it involves a learning component as well as elements affected by the school environment and infrastructure. It is, therefore, imperative to assess the knowledge, attitude, and practice of Secondary School girls in Ogbomoso on the subject of menstruation and menstrual hygiene.Methods: It was a cross-sectional study carried out between June and August 2016. A total of 447 eligible female adolescent secondary school students were selected by multistage sampling technique. A semi-structured facilitated self- administered questionnaire was used.Results: Majority (96.4%) of the adolescents have heard about menstruation before menarche while 55.9% had good knowledge of menstruation and menstrual hygiene. Despite the high level of knowledge, about 64% of the respondents describe their response to their first menses as scary, discomforting or emotionally disturbing. Also, only 25% of the respondents had good menstrual hygiene practice. Factors that were significantly associated with good knowledge of menstruation and menstrual hygiene in this study were older adolescent age (15-19 years) (χ2=47.68, P<0.0001); adolescents whose mothers have greater than secondary school education (χ2=42.52, P<0.0001); and those living with their parents (χ2=123.9 ,P=0.001) while only living with parents significantly influenced good practices.Conclusions: The study concluded that majority of the adolescents do not observe good hygienic practice during menstruation despite relatively good knowledge of menstruation and menstrual hygiene. Efforts have to be put into ensuring that adolescents are well educated on the meaning of menstruation, the importance of good hygienic practice both in schools and at home.