AIM: To investigate the content of serum microRNA-126 (miR-126) and its role in screening retinal endothelial injury and early diagnosis of proliferative diabetic retinopathy. METHODS: The study included 184 serum samples, 59 samples from healthy individuals, 44 samples from diabetes mellitus (DM) patients without diabetic retinopathy (NDR), 42 from non-proliferative diabetic retinopathy (NPDR) patients and 39 samples from proliferative diabetic retinopathy (PDR) patients. The expression of miR-126 was evaluated using a real-time quantitative polymerase chain reaction. RESULTS: The serum content of miR-126 declined as the damage degree in the retina. There was significant difference between the two retinopathy groups (P<0.001). No difference was observed in miR-126 content between healthy individuals and NDR patients (P>0.05). Receiver operating characteristic curve (ROC) analyses indicated that serum miR-126 had significant diagnostic value for PDR. It yielded an area under the curve (AUC) of ROC of 0.976 with 81.21% sensitivity and 90.34% specificity in discriminating PDR from healthy controls, and an AUC of ROC of 0.919 with 84.75% sensitivity and 94.41% specificity in discriminating NDR and NPDR from healthy controls. When the diagnostic threshold was greater than or equal to 8.43, there was an increase in the possibility of NPDR. When the content of miR-126 was less than or equal to 5.02, the possibility of the occurrence of PDR increased. CONCLUSION: Serum miR-126 can serve as a non-invasive biomarker for screening retinal endothelial injury and early diagnosis PDR.