This study evaluated the microbiological, nutritional, sensory, and in vitro biological properties of probiotic fermented milks enriched with 5% Prickly Pear Seed (PPS) flour. Different formulations were tested: plain fermented milk (Control), control with adjunct lactobacilli with probiotic traits (Lactiplantibacillus plantarum LPAL; Lacticaseibacillus casei BGP 93; Lacticaseibacillus rhamnosus LR B; Lacticaseibacillus paracasei LC4 P1), fermented milk with PPS, and fermented milk with PPS and adjunct lactobacilli. PPS flour did not affect the starters or the adjunct lactobacilli viability. Throughout 40 days of storage, probiotic levels remained above the minimum threshold required for health benefits. PPS supplementation increased the in vitro antioxidant activity of the fermented milk formulations only when combined with some of the adjunct lactobacilli. The formulation containing PPS and L. casei BGP 93 (PPS_BGP93) exhibited superior antioxidant performance. In HCT8 cell lines, PPS and PPS_BGP93 reduced Reactive Oxygen Species levels. PPS_BGP93 also upregulated IL-10 gene expression, although this response was not consistent with that found in the presence of LPS after 40 days of storage. PPS and PPS_BGP93 displayed the highest mineral levels and total fiber (around 3%), allowing the nutrition claims to be a source of phosphorus, magnesium, and fiber. A panel test (0–9 scores) revealed that although mouthfeel was impaired by the enrichment of PPS, the PPS-enriched formulations achieved scores around 6 for overall acceptability.