The uncertainty evaluation in the determination of inorganic arsenic in tassle filefish by using High Performance Liquid Chromatography-Inductively Coupled with Plasma Mass Spectrometry was investigated, based on the GB 5009.11 2014 “national food safety standards on the determination of total arsenic and inorganic arsenic in food”. A mathematical model was established, referring to CNAS-GL006 Guidelines for “the Assessment of uncertainty in Chemical Analysis and JJF 1059.1 2012 Evaluation and Expression of Measurement Uncertainty”. The precision and recovery of As (III) and As (V) in maomile fish were verified. Meanwhile, the sources of uncertainty measurement of the experiment were analyzed and evaluated. The results showed that the extended uncertainty of inorganic arsenic, i-As, was 0.007 with 95% confidence interval, when the content of inorganic arsenic in tassle filefish was 0.096 mg/kg. The main factors influencing this determination uncertainty were found to be the calibration curve and recovery rate.