Background: Low milk supply is frequently reported as a reason for exclusive breastfeeding cessation. Research aims: To determine the occurrence of, and the risk factors associated with, delayed onset of lactogenesis II among primiparas seen at a Baby-Friendly Hospital in Brazil. Method: We conducted a prospective longitudinal observational cohort study of 224 primiparas who had a singleton delivery. Data were first collected at the hospital. We assessed the onset of lactogenesis on day four postpartum, based on maternal reports of changes in breast fullness. Breastfeeding practices and Edinburgh Postnatal Depression Scale were evaluated on day seven postpartum. Using Poisson regression, we assessed significant factors associated with delayed onset of lactogenesis II. Results: Delayed lactogenesis II occurred in 18.8% ( n = 42) of participants and was significantly associated with alcohol drinking during pregnancy (IRR = 2.710, 95% CI [1.469, 4.996]); Edinburgh Postnatal Depression Scale scores ≥ 10 (IRR = 2.092, 95% CI [1.118, 3.916]), and the age of the mother (IRR: 1.081, 95% CI [1.039, 1.125]). Conclusion: Postpartum depression and alcohol ingestion during pregnancy may be associated with lactogenesis II delay, but more research is needed to elucidate the directionality of these relationships. Older mothers are at risk of delayed lactogenesis II onset. The frequency of delayed lactogenesis in this population is similar to the rates seen in previous Latin America studies and much lower than the ranges seen in North America, possibly because of the low proportion of obesity and severe gestational diabetes in this sample.