Prepartum supplementation with conjugated linoleic acid (CLA) may influence lipolysis and milk fat synthesis. The objective of this study was to examine the effect of prepartum CLA supplementation on lactation and metabolic health of dairy cows. Multiparous cows were enrolled in the study at −18 d prepartum, and randomly assigned 20 g/d of a mixture of trans-10,cis-12 and cis-9,trans-11 CLA (CLA n = 143; 100 g/d of Lutrell Pure; BASF) or an equivalent amount of saturated lipids as Control (Ctl n = 141; 75 g of Energy Booster 100; Milk Specialties Global). Treatments were top dressed individually to cows daily from enrollment to calving and all cows were offered the same ration. Blood samples were collected on the first day of supplementation, −10 d prepartum, and d 1, 7, 14, and 30 of lactation. Milk yield was recorded daily until 60 DIM and averaged weekly. A milk sample was obtained weekly for component analysis. Categorical data were analyzed by logistic regression (GLIMMIX, SAS) fitting a binary distribution response. Continuous variables were analyzed with the MIXED procedure of SAS. Models containing the fixed effect of treatment (GLIMMIX) or treatment, time, and treatment x time (MIXED) and random effects of block and cow(block × treatment). Prepartum treatment period was 16.1 ± 4.5 and 16.4 ± 4.3 for Ctl and CLA, respectively. Cows supplemented with CLA had increased milk protein yield (1.38 vs. 1.43 ± 0.019 kg, P = 0.01), and tended to have increased milk fat (1.94 vs. 2.00 ± 0.025 kg, P = 0.07) and milk yield (46.6 vs. 47.6 ± 0.45 kg, P = 0. 09), that resulted in greater energy content of milk (35.35 vs. 36.36 ± 0.379 Mcal/d, P = 0.03). CLA supplemented cows had a tendency for lower serum NEFA (0.28 vs. 0.23 ± 0.016 mEq/L, P = 0.06) and serum BHBA (0.96 vs. 0.80 ± 0.067 mM, P = 0.09), which resulted in decreased prevalence of hyperketonemia on d 14 postpartum (23.5% vs. 3.5% ± 5.79, P = 0.05). Body condition score change was not affected (P > 0.51) by treatment. There were no significant differences in other health disorders (P > 0.20). Prepartum supplementation of CLA improved lactation performance and metabolic health of dairy cows.