The results of epidemiological studies involving n-3 PUFA and polycystic ovary syndrome (PCOS) are scarce. This matched case–control study assessed the associations between n-3 PUFA and PCOS prevalence in 325 pairs of PCOS cases and healthy controls. Dietary information was assessed using a 102-item FFQ. Fatty acids in serum phospholipids were measured with a GC method. We found that n-3 PUFA in serum phospholipids were inversely associated with PCOS prevalence, including total, long-chain and individual PUFA (e.g. docosapentaenoic acid (DPA), EPA and DHA). Compared with the lowest tertile (T1), the adjusted OR and their 95% CI for the highest tertile (T3) were 0·63 (0·40, 0·93) for total n-3 PUFA, 0·60 (0·38, 0·92) for long-chain n-3 PUFA, 0·68 (0·45, 1·01) for DHA, 0·70 (0·45, 1·05) for EPA and 0·72 (0·45, 1·08) for DPA. For dietary intake of n-3 PUFA, significant inverse associations were found only for long-chain n-3 PUFA (P
trend = 0·001), EPA (P
trend = 0·047) and DHA (P
trend = 0·030). Both dietary and serum n-3 PUFA, mainly EPA and DPA, were negatively correlated with PCOS-related parameters, such as BMI, fasting insulin, total testosterone and high-sensitivity C-reactive protein, but positively correlated with follicle-stimulating hormone and sex hormone-binding globulin. These results indicated inverse associations between n-3 PUFA, especially long-chain n-3 PUFA, and PCOS prevalence. Higher intakes of n-3 PUFA might be considered a protective factor for PCOS among Chinese females.