Calcium is an important nutrient but also contributes to the texture, taste and functionality of most dairy products. The balance between micellar and free calcium in curd and serum phase during direct acidification of bovine milk was studied by changing organic acid type, pH and acidification temperature. Concentration of free calcium increased with decreasing pH and increasing temperature. Citric acid was more efficient than lactic acid at lowering free calcium concentration, while temperature effect was larger for acidification with lactic acid. These findings are rationalized by the stronger binding of calcium to citrate compared to lactate with an almost athermal binding for citrate and an endothermal binding for lactate. Curd yield decreased by decreasing pH as solubilization of colloidal calcium phosphate causes loss of minerals into the serum phase decreasing the concentration of both calcium and phosphate left in the curd, while acidification temperature had little effect on curd yield. These results are important to improve yield and tailor the properties and functionality of dairy products, such as for example Mozzarella cheese.