Introduction: Diabetic retinopathy is a common complication of diabetes and a leading cause of vision loss. The prevalence of DR increases with the duration of diabetes, poor glycemic control, and the presence of other systemic conditions such as hypertension and dyslipidemia. Serum C-peptide is a marker of endogenous insulin production and is commonly used to assess residual beta-cell function in patients with type 2 diabetes (T2D). Objectives: To examine the relationship between serum C-peptide levels and diabetic retinopathy in patients with type 2 diabetes, categorized according to their estimated glomerular filtration rate. Methods: A cross-sectional study was conducted in VINAYAKA MISSION’S KIRUPANANDA VARIYAR MEDICAL COLLEGE AND HOSPITALS, VINAYAKA MISSION’S RESEARCH FOUNDATION (DU), in the Department of General Medicine. The study was performed between September 2023 and July 2024. A total of 120 patients who had diabetes mellitus attending the OPD were included in the study. Blood samples were taken to measure serum C-peptide levels, fasting plasma glucose (FPG), and glycated hemoglobin (HbA1c). Serum C-peptide levels were measured using an electrochemiluminescence immunoassay. Also, eGFR was calculated using the CKD-EPI equation based on serum creatinine levels. Results: The majority of the participants had more than 10 years of diabetes (51.7%), while 48.3% had less than 10 years of diabetes. The mean HbA1c value was 8.2 ± 1.5, and the mean Serum C-peptide was 2.3 ± 1.1 ng/mL. The prevalence of diabetic retinopathy among the study participants was 45.0%. Conclusion: Serum C-peptide levels are significantly correlated with both diabetic retinopathy and kidney function. Lower C-peptide levels (reflecting worse beta-cell function) are associated with a higher risk of retinopathy and worse kidney function. © 2025 Elsevier B.V., All rights reserved.