Bovine mammary secretions were obtained during late lactation and early involution to determine if different methods of milk cessation influenced milk yield, composition, and in vitro growth of coliform mastitis pathogens. Cows (n=8/group) producing about 13kg of milk prior to experimentation were dried off by abrupt or intermittent milk cessation. An additional group was dried off by intermittent milk cessation and fed only hay during the last week of lactation. Cows milked intermittently produced significantly less milk during the last week of lactation than cows dried off by abrupt milk cessation. Mammary secretions from cows milked intermittently and fed only hay contained higher concentrations of somatic cells, lactoferrin, immunoglobulin G, and bovine serum albumin, a lower citrate:lactoferrin molar ratio, and were more inhibitory to in vitro growth of Escherichia coli and Klebsiella pneumoniae throughout most of the experimental period than mammary secretions from cows dried off by intermittent or abrupt milk cessation. Few differences in mammary secretion composition or in vitro growth of mastitis pathogens were observed between cows dried off by intermittent or abrupt milk cessation. Data suggest that growth of mastitis pathogens in mammary secretions may be related to natural protective factors, which can be manipulated by different methods of milk cessation.