Cow's milk allergy (CMA) is one of the most common food allergies in infancy, with prevalence estimates of 0.5-7.5% in high-income countries. Data from low- and middle-income regions remain limited, and the predominant immune mechanism (IgE or non-IgE mediated) may vary across populations. We aimed to determine the prevalence and clinical characteristics of CMA in infants from an urban, low-income Chilean population. A prospective cohort study was conducted at Padre Hurtado Hospital in Santiago, Chile. Healthy term newborns were recruited and followed for up to 12 months. Sociodemographic, perinatal data and parental atopy were recorded. Parents were contacted monthly to screen for CMA symptoms. Infants with ≥two symptoms underwent clinical evaluation, a 4-week cow's milk protein exclusion diet, and an open oral food challenge (OFC). Diagnosis followed international consensus guidelines. Of 552 enrolled infants (48% male), 27 were diagnosed with CMA, yielding a prevalence of 4.9% (95% CI 3.1-7.0%). All cases exhibited non-IgE-mediated symptoms, including vomiting, dermatitis, colic, and perianal erythema. CMA was diagnosed before 6 months of age in 74% of cases. At 12 months, 40% had developed oral tolerance. Sociodemographic and perinatal characteristics were similar between groups, but some self-reported parental atopic traits were more frequent in CMA cases. CMA prevalence in this Chilean cohort was comparable to that reported in high-income countries, with a predominance of non-IgE-mediated forms. These findings support the need for standardized diagnostic protocols, including OFC, in diverse populations. Future studies should explore long-term outcomes and risk factors in non-IgE-mediated CMA.