OBJECTIVE To identify milk component alterations that might be useful for detecting cows with rumen indigestion. DESIGN Prospective case-control study. ANIMALS 23 Holstein cows with rumen indigestion (cases) and 33 healthy cohorts (controls) from 1 herd. PROCEDURES Cases were defined as cows between 30 and 300 days postpartum with a > 10% decrease in milk yield for 2 consecutive milkings or > 20% decrease in milk yield from the 10-day rolling mean during any milking, abnormally decreased rumen motility, and no other abnormalities. Each case was matched with 2 healthy cows (controls) on the basis of pen, parity, days postpartum, and mean milk yield. Some cows were controls for multiple cases. All cows underwent a physical examination and collection of a rumen fluid sample for pH measurement at study enrollment. Individual-cow milk yield and milk component data were obtained for the 16 milkings before and after study enrollment. Rumen motility and pH and milk components were compared between cases and controls. RESULTS Rumen motility for cases was decreased from that of controls. Cases had an abrupt increase in milk fat percentage and the milk fat-to-lactose ratio during the 2 milkings immediately before diagnosis of rumen indigestion. Receiver operating characteristic analyses revealed that a 10% increase in the milk fat-to-lactose ratio had the highest combined sensitivity (57%) and specificity (85%) for identifying cows with rumen indigestion. CONCLUSIONS AND CLINICAL RELEVANCE Results indicated that a positive deviation in the milk fat-to-lactose ratio might be useful for identifying cows with rumen indigestion.