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Background: The associations of serum omega-6 (n−6) fatty acids with lipoprotein subclasses at the population level are uncertain. Objective: We aimed to examine associations between major n−6 fatty acids [ie, linoleic acid (LA, 18:2n−6) and arachidonic acid (AA, 20:4n−6)] and the lipoprotein subclasses VLDL, LDL, and HDL. Design: We conducted a cross-sectional study in 1098 participants using population-based data from US white, Japanese American, Japanese, and Korean men aged 40–49 y. Serum fatty acids were analyzed by capillary gas-liquid chromatography. Lipoprotein subclasses were measured by nuclear magnetic resonance spectroscopy. Multiple linear regression models as a function of each fatty acid were used after adjustment for age, population, body mass index, pack-years of smoking, alcohol consumption, diabetes, hypertension, and omega-3 (n−3) and trans fatty acids. Results: Serum LA was inversely associated with large VLDL (β = −0.62, P < 0.001), total LDL (β = −22.08, P < 0.001), and small LDL (β = −31.89, P < 0.001) particle concentrations and VLDL size (β = −0.72, P < 0.001). Serum LA was positively associated with large HDL particle concentration (β = 0.21, P < 0.001) and HDL size (β = 0.03, P < 0.001). The patterns of association of AA with large VLDL and large HDL particle concentrations were comparable with those of LA. Conclusions: At the population level, higher serum concentrations of LA were significantly associated with lower concentrations of total LDL particles. Higher serum concentrations of LA and AA were significantly associated with a lower concentration of large VLDL particles and a higher concentration of large HDL particles. These associations were consistent across the population groups. This trial was registered at clinicaltrials.gov as {"type":"clinical-trial","attrs":{"text":"NCT00069797","term_id":"NCT00069797"}}NCT00069797.