Background: Male partner involvement is an important and crucial determinant of prevention of mother to child transmission (PMTCT) of HIV. It creates an opportunity to reverse the transmission of HIV during pregnancy, labor, and breastfeeding. Thus, involving male partners during HIV screening of pregnant mothers at ANC is the key to fight against MTCT of HIV. Objective: This study was designed to assess the magnitude and factors associated with male partner's involvement on PMTCT service utilization among pregnant women who attended focused antenatal care (FANC) in Southern Ethiopia. Methods: An institutional-based cross-sectional study was conducted among 420 randomly selected pregnant women who enrolled in PMTCT service at ANC clinics. Pretested and structured self-administered questionnaires were used to collect the data. Multiple logistic regression analysis was used to determine the presence of statistically significant associations between the outcome variable and the independent variables with a p-value less than 0.05. Results: A total of 409 pregnant women who had ANC follow-up have participated in this study. The majority 160 (39.1%) of the participants were in the age group of 25-29 years. The magnitude of male involvement in PMTCT service was 129 (29.8%). Number of ANC visits (3rd visit (AOR=2.36, CI=1.09, 5.10), 4th visit (AOR=3.49, CI=1.65, 7.38), birthplace interest (AOR=3.01, CI=1.16, 7.84), awareness about partner monthly income (AOR=2.17, CI=1.15, 4.11), source of family saving scheme (partner (AOR=2.99, CI=1.39, 6.43), self (AOR=8.59, CI=3.92, 18.82), both (AOR=5.13, CI=2.21, 11.92), maternal perception about the importance of consulting partner before HIV testing (AOR=9.30, CI=2.65, 32.64), and kinds of partner support (psychological (AOR=0.08, CI=0.02, 0.29), financial (AOR=0.33, CI=0.17, 0.68) were found to be significantly associated with male involvement in PMTCT. Conclusion: This study found out that male partner involvement in PMTCT is low. Therefore, improving male partner involvement in PMTCT is recommended for improving maternal FANC service utilization and adherence with notification of their partner and provision of psychological and financial support.