Background: Kidney transplantation remains the gold standard therapy for paediatric endstage renal disease.Tacrolimus is the cornerstone immunosuppressant for preventing graft rejection.While peripheral venipuncture (VP) is the reference method for herapeutic drug monitoring (TDM), it proves challenging in paediatric patients due to compromised vascular access and limited cooperation. Repeated VP heightens the risk of trauma, haematoma, and infection. Conversely , central venous catheters (CVCs), routinely placed for post-operative care, offer a practical alternative to improve patient compliance. Objective: This study aims to prospectively compare the consistency and reliability of Tacrolimus levels in blood samples obtained from CVC versus VP in pediatric kidney transplant patients, providing an evidence base for optimising clinical sampling practices. Methods: This prospective study included pediatric kidney transplant patients with central catheters free of intravenous Tacrolimus. Paired samples were collected daily for seven days via peripheral venipuncture and central catheter using a standardised “mixed push-pull” protocol. Pearson correlation and Bland-Altman analyses quantified the linear relationship and mean difference between the measures.Ethical Review Approval Number: Fu'er Ethical Review (4) 374. Results: Analysis of 105 paired samples revealed no significant difference in mean Tacrolimus levels between the CVC and VP groups (P > 0.05). A robust positive correlation was observed between the methods (r= 0.969, P < 0.001). Bland-Altman analysis demonstrated excellent agreement with a minimal mean bias of 0.276 ng/mL (95% CI: -2.637 to 3.189). Linear mixedeffects modelling confirmed high consistency between the measures at the overall mean level. No complications, such as catheter occlusion or infection, were observed.