Research suggests that conjugated linoleic acid (CLA) may inhibit atherosclerosis, but there are contradictory results in different animal models fed heterogeneous mixtures of CLA isomers. This study addressed the hypothesis that the individual CLA isomers may exert different atherogenic properties. ApoE(-/-) mice were fed isocaloric, isonitrogenous westernized diets containing 0.15% cholesterol and enriched with 1% (w/w) cis-9,trans-11-CLA (c9,t11-CLA), trans-10,cis-12-CLA (t10,c12-CLA) or linoleic acid (control diet) for 12 weeks. At the end of the dietary intervention, the effects of CLA isomers on the development of atherosclerotic vascular lesions, lipid metabolism, inflammation and oxidative stress were assessed. The t10,c12-CLA diet had a profound pro-atherogenic effect, whereas c9,t11-CLA impeded the development of atherosclerosis. En face aortic lesion assessment showed more dorsal and lumbar extensions presenting atherosclerotic foci after the t10,c12-CLA diet. Furthermore, animals fed t10,c12-CLA had pronounced hyperlipidemia, higher 8-iso-prostaglandin F(2alpha) levels, higher vulnerable atherosclerotic plaque with a lower smooth muscle and fibre contents and higher macrophage content and activation, assayed as plasma chitotriosidase compared to the control or c9,t11-CLA dietary groups. Plasma chitotriosidase activity was more closely associated with the extent of the plaque than with MOMA staining or than monocyte chemoattractant protein-1 levels. Our results demonstrate that CLA isomers differentially modulate the development of atherosclerosis, c9,t11-CLA impedes, whereas t10,c12-CLA promotes atherosclerosis. These opposing effects may be ascribed to divergent effects on lipid, oxidative, inflammatory and fibro muscular components of this pathology. Plasma chitotriosidase is a better indicator of dietary fat interventions that alter plaque monocyte activity in this murine model.