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The health benefits of PUFA are well established. There is no valid tool or complete fatty acid database to assess PUFA intake in New Zealand (NZ). This study aimed to develop, validate, and test the reproducibility of a NZ-specific PUFA FFQ. A semiquantitative NZ PUFA FFQ was developed based on a validated Australian PUFA FFQ. The Australian fatty acid database was adapted to include NZ-specific data for major PUFA sources. Healthy participants from Auckland, NZ (n = 48) provided fasting blood samples for erythrocyte PUFA analysis, completed the NZ PUFA FFQ and a 3-d weighed food record (WFR), and repeated the NZ PUFA FFQ 3 mo later (n = 42). Relative validity was evaluated by assessing the triangular relationship among the NZ PUFA FFQ, WFR, and erythrocyte PUFA using the methods of triads [EPA, DHA, total omega-3 (n-3) long-chain (LC) PUFA only] and by comparing, correlating, cross-classifying into quintiles and assessing agreement using Bland-Altman plots of intakes between the NZ PUFA FFQ and WFR. Reproducibility was assessed by comparing and correlating intakes between repeat administrations of the NZ PUFA FFQ. The NZ PUFA FFQ effectively estimated EPA [ρ(QT) = 0.72 (95% CI: 0.49, 0.89)], DHA [ρ(QT) = 0.72 (95% CI: 0.53, 0.95)], and total (n-3) LCPUFA [ρ(QT) = 0.68 (95% CI: 0.47, 0.89)] intakes and was comparable with the WFR for other PUFA except docosapentaenoic acid. Repeated implementation of the NZ PUFA FFQ showed agreement for PUFA intakes. The NZ PUFA FFQ is a valid and reliable tool to measure PUFA intake in healthy NZ adults.