This study investigated the differences in absorption of infant formula with different levels of α-lactalbumin (α-La) and β-casein (β-CN), and how α-La and β-CN (α+β) affect mineral absorption using the Caco-2 cell model. Compared to the control group, the free amino acid content transported to the basal side significantly increased in the fortification of α-La and β-CN, particularly in the α+β (high) group, which reached 11.343 μmol/L. Furthermore, fortification of α-La and β-CN upregulated the expression of the amino acid transporter SLC6A14 and the oligopeptide transporter PEPT1, in particular the expression of SLC6A14 in the α+β (high) group was 103% of that of the control group. In addition, fortification of α-La and β-CN enhanced the absorption of Ca, Mg, and Zn. The absorption rates of Ca, Mg, and Zn in α+β (high) group increased to 31.27%, 13.66%, and 58.59%, respectively, of which α-La contributed the most on Ca absorption. In conclusion, α-La and β-CN fortification not only increased the absorption of amino acids and peptides but also promoted the absorption of minerals, which provide evidence for the benefits of adjustment of α-La and β-CN in the infant formula.