Background: Contradicting results have been published regarding the effect of conjugated linoleic acid (CLA) on insulin resistance. However, only a few studies have used the euglycemic hyperinsulinemic clamp method, which is considered the standard for measuring insulin resistance. Objective: To evaluate if CLA as a mixture of the main isomers trans-10 cis-12 and cis-9 trans-11 affects the insulin resistance in healthy overweight and obese male and female adults. Design: The main study was a randomized, double-blind, placebo-controlled trial with change in body composition as primary end point comprising 118 subjects receiving supplementation with either placebo (olive oil) or CLA (Clarinol) for 6 months. A sub-population of 49 subjects agreed additionally to participate in an euglycemic hyperinsulinemic clamp study at baseline and after 6 months of supplementation with study drug. The primary outcome was the change in glucose uptake (M) as measured by the hyperinsulinemic euglycemic glucose clamp method. Secondary outcomes were the correlates between insulin resistance and changes in body composition or blood chemistry parameters. Forty-one subjects completed the clamp test at both time points. Results: The median M of the CLA group was 11.0 mg min À1 lean body mass (lbm) À1 (n ¼ 24) at baseline, 10.3 mg min À1 lbm À1 (n ¼ 24) after 6 months, and the median difference was þ 0.21 mg min À1 lbm À1 (n ¼ 24). The median M of placebo group was 8.4 mg min À1 lbm À1 at baseline and 9.3 mg min À1 lbm À1 after 6 months and the median difference was À0.22 mg min À1 lbm À1 (n ¼ 17). No significant (Po0.05) differences were found within groups or between groups. Likewise, the glucose uptake insulin concentration ratio during clamp (M/I) was independent of treatment and time. Homeostasis model assessment (HOMA) and quantitative insulin sensitivity check index derived from fasting glucose and insulin were also independent of treatment and time, and HOMA for the clamp population (n ¼ 49) corresponded well with HOMA for the per protocol population (n ¼ 83). Correlation analysis showed that changes in M were inversely correlated to changes in glucohemoglobin (P ¼ 0.002), but did not correlate with changes in either glucose, insulin, insulin c-peptide, leptin, adiponectin or percent body fat. Conclusions: CLA does not affect glucose metabolism or insulin sensitivity in a population of overweight or obese volunteers.