The influence of dietary fats on in vitro lipid absorption by bovine intestine was studied in 14 calves. Holstein bull calves were fed for 16 wk five liquid diets containing skim milk plus either 3.5% milk fat, 3.5% tallow, 3.5% tallow and .2% cholesterol, 7.0% tallow, or 7.0% tallow and .2% cholesterol. Uptake of oleic acid or triolein by everted jejunal or ileal sacs was measured after incubation for 30 min at 37 degrees C in pH 7.4 micellar solutions containing tritium-labeled oleic acid or tritium-labeled triolein. Lipids were extracted from homogenates of sacs and separated into lipid classes by thin layer chromatography. Equal amounts of oleic acid were taken up by jejunal or ileal sacs. Triolein uptake was less than oleic acid uptake, but uptakes by jejunal or ileal sacs did not differ. Oleic acid incorporation into triglyceride was three to four times greater in intestinal sacs from milk fat-fed calves than in sacs from calves fed either 3.5 or 7% tallow. Oleic acid incorporation in intestinal sacs from calves fed cholesterol as well as tallow was equal to that in milk fat-fed calves. Intestinal uptake and metabolism of oleic acid proceeded faster when calves were fed milk fat than when fed tallow. The lower intestinal incorporation with tallow increased if cholesterol was fed, suggesting that cholesterol either stimulated absorption or increased esterification.