Background: Exclusive breast feeding (EBF) of Human Immune Virus (HIV) exposed infants for the first six months is strongly recommended and vital for protecting them against common childhood illnesses including diarrhea and pneumonia. Despite its benefit, EBF practice is low in developing countries including Ethiopia. There is a paucity of evidence for factors associated with EBF among HIV positive mothers. Objectives: To assess exclusive breast feeding practice and its associated factors among HIV positive mothers attending Prevention of mother to child transmission and Anti-retroviral therapy clinics in public Health facilities of Debre Birhan town Amhara National Regional State, Ethiopia from February 01, 2020 to Apr 30, 2020Methods: Facility based convergent mixed methods design was used to conduct the study in public Health facilities of Debre Birhan town. Structured interviewer-administered questionnaires were used to collect data from 432 participants selected by systematic random sampling technique. Epi info version 7 and SPSS version 20 were used for data entry and analysis respectively. Descriptive and inferential statistics were computed during the analysis. Two focus group discussions and 25 in-depth interviews and observational checklist were held to collect data on barriers for exclusive breast feeding practice and thematic analysis was used to identify important insights on the study topic. Bivariate and multivariate logistic regression were conducted to select candidate variable and determine adjusted effect of independent variables respectively using p-value less than 0.05. The output from multivariate logistic regression were reported with 95% confidence interval.Results: The prevalence of exclusive breast feeding, mixed feeding and exclusive formula feeding practice were 89.8%, 6.9% and 3.2% respectively. Having information about EBF (No=0.02, 95% CI; 0.01, 0.12), time taking from home to workplace (less than 30 min=4.96, 95% CI; 1.17, 20.95), disclosing HIV status (No= 0.09, 95% CI; 0.02, 0.37), place of residence (urban=5.37, 95% CI; 1.12, 25.77), living with mother/mother in-law (No=6.03, 95% CI; 1.26, 28.86), knowledge about EBF (poor knowledge=0.06, 95% CI; 0.01, 0.34) were main factors for exclusive breast feeding practice. Poor counseling, non-disclosure of HIV status and pressure from mother/mother in-law were the commonly raised reasons by FGD and IDI participants for non-exclusive breast feeding practice. Conclusions: Majority of HIV positive mothers were practicing exclusive breast feeding for the first six month. Strengthening information dissemination, providing quality health services and empowering mothers through integrated interventions help to promote exclusive breast feeding practice among mothers living with HIV.