Background: Regional citrate anticoagulation (RCA) has emerged as a treatment modality that reduces bleeding risk and filter clotting. With initial experience of using RCA with continuous renal replacement therapy (CRRT), we have formulated a working protocol based on published literature. Objective: The study aimed to evaluate the protocol for routine use of RCA during CRRT requiring anticoagulation and evaluation of filter life. Methodology: It is a single-center, open-label, prospective, non-randomized, non-interventional, single-arm, observational study conducted at a tertiary care hospital between September 2022 and July 2023. All adult patients with acute kidney injury (AKI) or hyperammonemia requiring CRRT and necessitating the use of anticoagulation were enrolled in the study. The study used Prisma Flex M100 AN 69 dialyzer on Prisma Flex (Baxter) CRRT machines during continuous venovenous hemodiafiltration (CVVHDF). The targeted CRRT dose in all the study patients was 25–30 mL/kg/hour. Based on the published literature, we have developed a working protocol (Appendix 1) for managing patients on CRRT using RCA. Results: A total of 159 patients were analyzed for the study. The median [interquartile range (IQR)] filter life using RCA was 30 (12–55) hours. Filter clotting was observed in 33.3% of patients. Citrate accumulation was present in 52.25% of patients, but no CRRT was discontinued as citrate accumulation resolved after following the corrective steps in the protocol. None of the patients had citrate toxicity. Chronic liver disease (CLD) (p ≤ 0.001) and those who were post-living donor liver transplant recipients (p = 0.004) had a statistically significant increase in citrate accumulation. Also, patients who had higher lactate at baseline (6 hours post-CRRT initiation), had a higher chance of citrate accumulation. Conclusion: Our RCA protocol provides a safe approach to regional anticoagulation during CRRT in critically ill patients.