Background: Recently, peroxiredoxin3 (PRDX3) was identified as a novel molecular marker for the progression of hepatocellular carcinoma (HCC). However, its potential clinical application as a serum marker for the early diagnosis and prognosis of HCC has not been investigated. Methods: PRDX3, alpha-fetaprotein (AFP), and other biochemical parameters were measured in serum samples from 297 Chinese patients, including 96 with HCC, 98 with liver cirrhosis (LC), and 103 healthy controls (HCs). Correlations between serum PRDX3 expression and clinicopathological variables and the relationship between serum PRDX3 expression and prognosis were analyzed. Results: Serum PRDX3 was significantly higher in HCC patients than in the LC and HC groups. The sensitivity and specificity of serum PRDX3 for the diagnosis of HCC were 85.9% and 75.3%, respectively, at a cutoff of 153.26 ng/mL, and the area under the curve was 0.865. Moreover, serum PRDX3 expression was strongly associated with AFP level, tumor diameter, TNM stage, and portal vein invasion. Kaplan-Meier curve analysis revealed that HCC patients with high serum PRDX3 expression had a shorter median survival time than those with low PRDX3 expression. Moreover, serum PRDX3 expression was an independent risk factor for overall survival. The inverse correlation between serum PRDX3 and patient survival remained significant in patients with early-stage HCC and in those with normal serum AFP levels. Conclusions: Serum PRDX3 can be used as a noninvasive biomarker for the diagnosis and/or prognosis of HCC.