Conjugated linoleic acid (CLA) supplements have typically been comprised of 4 isomers (trans-8, cis-10; cis-9, trans-11; trans-10, cis-12; and cis-11, trans-13 CLA). Abomasal infusion of pure isomers has shown that trans-10, cis-12 CLA is a potent inhibitor of milk-fat synthesis, whereas cis-9, trans-11 CLA has no effect. However, there appear to be additional fatty acids that inhibit milk-fat synthesis, and the objective of this study was to investigate the effects of additional CLA isomers present in CLA supplements. Four rumen fistulated Holstein cows (141+/-8 DIM, mean+/-SE) were randomly assigned in a 4 x 4 Latin square experiment. Treatments were abomasal infusion of (1) skim milk (negative control), (2) trans-10, cis-12 CLA supplement (positive control), (3) trans-8, cis-10 CLA supplement, and (4) cis-11, trans-13 CLA supplement. Treatments 2 through 4 were targeted to provide 4 g/d of the CLA isomer of interest. The trans-8, cis-10 CLA supplement had no effect on milk-fat yield, whereas the trans-10, cis-12 CLA supplement reduced milk-fat yield by 35%. The cis-11, trans-13 CLA supplement contained some trans-10, cis-12 CLA, and when data were compared to the positive control treatment group, it was obvious that cis-11, trans-13 CLA also had no effect on milk-fat synthesis. Milk-fat content of specific CLA isomers was significantly elevated within respective treatment groups. Milk yield, DMI, and milk protein yield were unaffected by treatment. Overall, trans-10, cis-12 CLA reduced milk-fat synthesis, whereas the other major isomers present in CLA supplements (trans-8, cis-10 CLA and cis-11, trans-13 CLA) had no effect.