INTRODUCTION: Tenofovir disoproxil fumarate (TDF) is a third-generation nucleoside analogue commonly used as a first-line therapy for chronic hepatitis B virus (HBV) infection. However, it is associated with nephrotoxicity, which can occur through mechanisms such as mitochondrial deoxyribonucleic acid (DNA) depletion and damage to renal tubules. This nephrotoxic effect typically manifests as a mild increase in serum creatinine and a decrease in serum phosphate, usually within 4-12 months after starting treatment. Recent studies suggest that novel biomarkers, such as urinary neutrophil gelatinase-associated lipocalin (NGAL) and cystatin C, may predict acute kidney injury (AKI) earlier and more accurately than traditional markers like serum creatinine. In line with this, we investigated NGAL and cystatin C levels in HBV patients receiving TDF therapy to assess their potential in monitoring kidney function. MATERIALS AND METHODS: The study included 350 cases in total. Each participant underwent a thorough assessment, including a detailed medical history, clinical examination, and routine blood tests, as outlined in the study's working proforma. Based on the above details, 226 cases fulfilling the inclusion criteria were enrolled for the second step of the study, where a 5 mL urine sample from each case was taken and sent to the pathology laboratory for estimation of cystatin C and NGAL by the ELISA method with the help of a kit. RESULTS: The average age of the participants was 37.93 years, with 124 individuals falling within the 18-35-year age-group. After accounting for confounding factors, 87 cases were identified, predominantly young males, who exhibited elevated levels of both urinary NGAL and cystatin C. CONCLUSION: In this cross-sectional study, after controlling for confounding factors, we observed that TDF treatment was linked to elevated levels of urinary NGAL and cystatin C in 87 (38.4%) of the cases. Notably, increased levels of these biomarkers were also found in the younger age-group (18-35 years). These findings suggest the need for further prospective studies with larger sample sizes to better understand the direct impact of TDF on kidney function in hepatitis B patients.