Ovarian high-grade serous carcinoma (HGSC) and low-grade serous carcinoma (LGSC) are distinct gynecologic neoplasms with diverse pathogenesis and characteristic features. They respond differently to same modalities of treatment protocol and have dissimilar prognosis. Thus, it is essential to obtain accurate differential diagnosis of HGSC and LGSC prior to clinical treatment. In the present study, mRNA expression profiles were generated from 5 HGSC and 6 LGSC specimen using HTSeq, and 699 differentially expressed genes (>2-fold difference) were identified using the DESeq R package. Dendrograms produced by unsupervised hierarchical clustering completely distinguished HGSC from LGSC. Among differentially expressed genes between HGSC and LGSC, anterior gradient homolog 3 (AGR3) was highly upregulated in LGSC compared to HGSC, which was validated by reverse transcription‑quantitative polymerase chain reaction and western blotting. Then, anti‑tumor protein 53 (TP53) and anti-AGR3 immunohistochemistry were performed on 145 HGSC and 30 LGSC samples. Consistent with previous studies, abnormal expression of TP53 (0 or ≥75% positive expression) was observed in 87.6% of HGSC and 13.3% of LGSC samples. Positive staining of AGR3 had a sensitivity of 80.0% and specificity of 89.7% for LGSC. TP53 and AGR3 were both efficient in distinguishing HGSC from LGSC (P<0.001). Receiver operating characteristic analysis revealed a similar area under the curve for AGR3 (0.848) and TP53 (0.871). Through combination of the two markers (TP53 wild‑type pattern and AGR3‑positive expression), the accuracy of differential diagnosis was up to 93.1%. These findings provide compelling evidence that differential diagnosis of HGSC and LGSC can be improved by combined application of these two markers on the basis of conventional histopathological diagnosis.