Objectives: Amyloid diseases are a rare class of disorders that have gained significant attention for therapeutic advancements. Among the key proteins implicated, transthyretin plays a central role in the progression and clinical symptoms of these diseases. Tafamidis, an approved therapeutic agent, has demonstrated considerable efficacy in mitigating symptoms associated with transthyretin-related amyloid conditions. Despite its clinical significance, robust bioanalytical methodologies for the accurate quantification of Tafamidis in biological matrices remain limited. Materials and Methods: This study focuses on the development and validation of a precise bioanalytical method to quantify Tafamidis in human plasma using Liquid Chromatography-Tandem Mass Spectrometry (LC-MS/MS). The method was calibrated over a range of 5-4000 ng/mL, with chromatographic separation performed using an isocratic mobile phase on a Gemini NX C18 column. Mass transitions were optimized at 307.400 → 262.400 for Tafamidis and 313.900 → 270.000 for Tafamidis-13C6. A liquid-liquid extraction approach ensured efficient recovery of Tafamidis from plasma. Results and Discussion: Validated per USFDA regulatory guidelines, the method demonstrated excellent precision, accuracy, and stability, with correlation coefficients exceeding 0.99 and recovery rates reaching 70%. Stability was confirmed for up to 10 days in matrix and aqueous samples. Conclusion: This robust, validated method offers high reliability and suitability for analytical applications and quantitative clinical investigations involving Tafamidis. © Author (s) 2026 Distributed under Creative Commons CC-BY 4.0.