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Abstract Background Mentor mothers provide psychosocial and other support to pregnant and post-partum women living with HIV (WLHIV), which has been shown to enhance maternal-infant outcomes in the prevention of mother-to-child transmission of HIV (PMTCT). Our objective was to assess the acceptability of mentor mothers as a PMTCT intervention, and to explore opinions on mentor mother program composition and delivery among stakeholders in North-Central Nigeria. Methods We conducted nine focus group discussions and 31 in-depth interviews with 118 participants, including WLHIV, pregnant women, male partners, health workers, traditional birth attendants, community leaders, PMTCT program implementers, and policymakers. Participants were purposively recruited from health facilities and surrounding communities in the Federal Capital Territory and Nasarawa State. Transcripts were manually analysed using a Grounded Theory approach, where theory was derived from the data collected. Results Most participants were female (n = 78, 67%), and married (n = 110, 94%). All participant groups found mentor mothers acceptable as women providing care to pregnant and postpartum women, and as WLHIV supporting other WLHIV. Mentor mothers were uniquely relatable as role models for WLHIV because they were women, living with HIV, and had achieved an HIV-negative status for their HIV-exposed infants. Mentor mothers were recognized as playing major roles in maternal health education, HIV treatment initiation, adherence, and retention, HIV prevention for male partners and infants, and couple HIV disclosure. Most WLHIV preferred to receive mentor mothers’ services at health facilities rather than at home, due to concerns about HIV-related stigma and discrimination through association with mentor mothers. Key mentor mother needs were identified as training, remuneration, and validation as lay health workers. Conclusions Mentor mothers are an acceptable PMTCT intervention among stakeholders in North-Central Nigeria. However, stigma and discrimination for both mentor mothers and their clients remain a critical challenge, and mentor mother needs such as training, pay, and a sustainably supported niche in health systems require focused attention. Trial registration Clinicaltrials.gov registration number ( NCT 01936753 ), registered on September 3, 2013 (retrospectively registered).