The relationship of the estrous cycle to milk composition was examined in 15 Holstein cows in first lactation which were approximately 45 days postpartum. Composite milk samples were collected twice daily and analyzed for progesterone, fat, protein, total solids, sodium, potassium, magnesium, calcium, and somatic cells. Milk progesterone was minimal (less than ng/ml) from day -1 to day 2 (day 0 = estrus) of the estrous cycle and then increased at a relatively constant rate through day 15. Milk yield and sodium and magnesium concentrations of the milk varied with days of the estrous cycle. Mean milk yield was highest and sodium and magnesium concentrations were lowest on day 1. However, sodium concentration of the milk was the only component that varied significantly during the 3 days centered on estrus (days -1, 0, and 1). No other milk component changed significantly during the estrous cycle. Although milk yield and composition varied during the estrous cycle, none of these components appeared to be a practical indicator of estrus.