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To observe the performance of human epididymis protein 4 (HE4), aldehyde dehydrogenase 1 (ALDH1) and adhesion molecule CD44 in differential diagnosis of benign and malignant ovarian tumors. 100 patients with ovarian carcinoma and 100 patients with benign ovarian tumor who were admitted to the hospital from January 2020 to May 2022 were selected, and included in the malignant group and the benign group. At the same time, 100 healthy women were selected as the control group. The expression of HE4 and ALDH1 in serum and CD44 in tissues of the three groups was compared. The value of the three parameters in differential diagnosis of benign and malignant ovarian tumors was analyzed using the receiver operating characteristic (ROC) curve. The levels of HE4 and ALDH1, and CD44 positive expression rate decreased in order from the malignant group, the benign group to the control group (P<0.05). For the malignant group, the levels of HE4 and ALDH1, and CD44 positive expression rate were higher in patients with clinical stage III-IV and lymph node metastasis than those with clinical stage I-II and without lymph node metastasis (P<0.05). Spesrman correlation analysis found that the levels of HE4 and ALDH1, and CD44 positive expression rate were positively correlated with clinical stage and lymph node metastasis in patients with ovarian carcinoma (P<0.05). ROC curve analysis results showed that the area under the curve (AUC) values of HE4, ALDH1, and CD44 in differential diagnosis of benign and malignant ovarian tumors were 0.837, 0.768, and 0.610, respectively. The AUC and sensitivity of combined diagnosis with the three indicators were higher than those of single diagnosis (P<0.05). The expression of HE4 and ALDH1 in serum and CD44 in tissues of patients with malignant ovarian tumors is high. The expression level is closely related to clinical stage and lymph node metastasis of ovarian carcinoma. The combination of HE4, ALDH1 and CD44 is of high value in the differential diagnosis of benign and malignant ovarian tumors. [ABSTRACT FROM AUTHOR]