Background Despite the progress made so far in reducing mother-to-child transmission (MTCT), Nigeria still contributes significantly to the global burden of new pediatric HIV infections. The elimination target for MTCT has not been reached and the decline in new infections among all Global Plan countries from 2009 to 2015 was lowest in Nigeria. This qualitative study explores the barriers to uptake of prevention of mother-to-child transmission (PMTCT) intervention in Kano, the second most populous state in Nigeria. Methods Key informant interviews (KIIs) were conducted among twelve stakeholders who were purposively selected based on their knowledge and involvement in PMTCT program activities in the state. The KII guide explored the status and challenges of PMTCT uptake in Kano state. Qualitative data analysis was managed using NVIVO 11 software and themes were analyzed using thematic analysis. Results We found that the key barriers to uptake of PMTCT identified by stakeholders cut across the domains of the socio-ecological model. These include-fear of stigma associated with being seen accessing HIV related services, low male partner involvement, socio-cultural beliefs about the dangers of hospital-based delivery, poor attitude of health workers, distance/cost to facilities, issues with availability of HIV test kits and poor organization of health services. Conclusion The implementation of effective PMTCT programs would require innovative strategies that leverage improvement of Antenatal care (ANC) uptake as an entry point for PMTCT. In addition, sustaining engagement in care requires creating a supportive stigma-free environment in the community as well as spousal support to ensure women can navigate the socio-cultural barriers that limit access to health services.