The objective of this study was to investigate the expression of thyroid peroxidase (TPO) in papillary thyroid carcinoma (PTC) and to preliminarily investigate its value as a marker of lymph node metastasis and recurrence in patients with PTC. Clinical data of PTC patients and TPO expression were collected from The Cancer Genome Atlas (TCGA) database for analysis. We recruited 230 consecutive PTC patients from the Department of Thyroid Surgery of the First Affiliated Hospital of Kunming Medical University, collected their clinicopathological data, and also performed immunohistochemical analysis of TPO expression on their thyroid specimens to validate the results of bioinformatics analysis. In addition, the construction of protein–protein interaction networks was performed too. Functional enrichment analysis and immuno-infiltration analysis characterized the pathways in which TPO genes may be involved. Data mining based on the TCGA database showed that TPO expression in PTC tissues was significantly lower than in paired normal thyroid tissues. The expression level of TPO in PTC tissues correlated with tumor lymph node metastasis and recurrence. Follow-up data from our center also validated the difference in TPO expression and its relationship with lymph node metastasis in PTC patients. Functional enrichment analysis showed that TPO function was significantly associated with signaling pathways related to amino acid metabolism, gene expression regulation and tumorigenesis. TPO expression was also significantly associated with immune infiltration. Our study showed that reduced TPO expression was significantly associated with lymph node metastasis and recurrence in patients with PTC, and we validated this result in our central cohort. These data suggest that TPO may serve as a prognostic indicator for PTC.