Purpose: To explore serum chemokine ligand 12 (C-X-C chemokine ligand 12, CXCL12) and chemokine ligand 16 (C-X-C chemokine ligand 16) The correlation between CXCL16 and poor postoperative prognosis in patients with Non-small cell lung cancer (NSCLC). Method: 100 NSCLC patients admitted to our hospital from February 2022 to June 2023 were selected as the research subjects. All patients have undergone radical resection treatment, and the levels of serum CXCL12 and CXCL16 were detected before the operation. The patients were followed up for 12 months after the operation. According to the different prognosis of the patients during the follow-up period, they were divided into the good prognosis group and the poor prognosis group. Collect and compare the baseline data of the two groups and the levels of serum CXCL12 and CXCL16. To analyze the correlation between serum CXCL12 and CXCL16 and the poor postoperative prognosis of NSCLC patients. Result: Among 100 NSCLC patients, 20 cases had recurrence and metastasis during the 1-year postoperative follow-up. They were included in the poor prognosis group, and the incidence of poor prognosis was 20.00% (20/100). The remaining 80 patients were included in the good prognosis group. There were no significant differences in baseline data such as gender, age, body mass index, smoking history, underlying diseases, lesion side, pathological type, tumor diameter, operation time, intraoperative blood loss, treatment plan, and occurrence of postoperative complications among patients with different prognostics (P>0.05). The TNM tumor stage, serum CXCL12 and CXCL16 levels in the poor prognosis group were significantly higher than those in the control group, and the degree of differentiation was significantly lower than that in the control group (P < 0.05). The results of COX regression analysis revealed that serum CXCL12 and CXCL16 were associated with poor postoperative prognosis in NSCLC patients (P < 0.05). Through dot two-column correlation analysis, both serum CXCL12 and CXCL16 were positively correlated with poor postoperative prognosis in NSCLC patients (r > 0, P < 0.05). The Receiver Operating Characteristic (ROC) curve was plotted. The results showed that the combined value of serum CXCL12 and CXCL16 levels in predicting poor postoperative prognosis in NSCLC patients was higher. Conclusion: Serum CXCL12 and CXCL16 are closely related to the occurrence of poor postoperative prognosis in NSCLC patients. Combined detection can predict the risk of poor postoperative prognosis in NSCLC patients.