Cheddar cheese was made from milk collected from two groups of cows milked either two or three times daily during early, mid, and late lactation. Milk from cows in late lactation had lower casein as a percentage of true protein and a higher acid degree value than did milk from cows in early lactation. Milk from cows milked three times daily had lower concentrations of milk fat and casein and higher acid degree values than did milk from cows milked twice daily, and thus this milk would be expected to result in decreased cheese yield. Cheese composition was not affected by milking frequency. Stage of lactation effects on cheese composition were confined to differences in salt content and a trend for higher moisture in cheese made from milk of cows in late lactation. Stage of lactation influenced the pH and degradation of alpha s-casein in cheese during aging. Fat and protein losses in whey at draining were higher for milk from cows in late lactation than from milk from cows in early lactation. The typical differences in fatty acid composition of milk from cows in early lactation that cause lower melting point may have caused higher fat loss in press whey. Fat loss in whey at draining was higher in cheese made from milk from cows milked three times daily than in cheese made from milk from cows milked twice daily, but the protein loss was not influenced. The ADV of milk was positively correlated to the fat loss in whey. Lower recoveries of fat and protein in cheese from milk of cows in late lactation were observed and may cause small but economically important decreases in cheese yield. Low SCC of milk from cows in late lactation may have minimized the changes in cheese composition and yield from stage of lactation.