The adrenal medulla is the inner part of adrenal glands located above each kidney, that produces catecholamines. Neuroblastoma and pheochromocytoma are the most prevalent malignancies of the adrenal medulla. Quantitative diagnosis of urinary catecholamines using HPLC-coupled Mass detectors is the current method for the diagnosis of neuroblastoma and pheochromocytoma. There are two major problems with this approach, (i) Because the catecholamines concentrations have short half-life (10-100 s), a series of urine tests must be performed throughout 24hr, detecting each catecholamine separately, is inconvenient and time-consuming; (ii) mass detectors are expensive, bulky, and require highly skilled personal. Vanillylmandelic (VMA), and homavanillic acid (HVA) are the by-products of catecholamines and are emerging alternative biomarker for catecholamines due to their high stability. Here, we developed a rapid, sensitive, miniaturized, and cheaper sensing platform for simultaneous quantifications of dopamine (DA), VMA, and HVA, with the aid of iron single-atom catalysts (Fe-SACs), based electrochemical sensor. SACs are atomically distributed metal atoms that have a maximum atomic utility rate of nearly 100%, compared to 30% for traditional metal nanoparticles. MXene sheets are employed to stabilize Fe-SACs, where, the exposed lone pairs of MXene serve as sites covalently linking high-energy single Fe atoms. MXene/Fe-SACs were synthesized by treating Ti3C2TxMXene with Iron chloride via freeze-drying followed by annealing. The successful formation of the material was verified by state-of-the-art characterizations. The MXene/Fe-SACs show superior electrocatalytic performance to the commonly used Fe- nanomaterials. Then, it was coated on the electrode surface and used to analyze DA, VMA, and HVA simultaneously via cyclic voltammetry (CV) and square-wave voltammetry (SWV). Under optimized conditions, the MXene/Fe-SACs electrochemical sensor showed detection limits as low as 1 nM and a linear range between 1 nM-100 μM for DA, LOD of 5 nM & linear range of 10 nM-100 μM VMA, and LOD of 10 nM & linear range of 20 nM-100 μM HAV. The method proved successful in detecting biomarkers in (spiked) synthetic urine and human serum. Furthermore, the method was successfully demonstrated in the determination of DA release from PC12 live cells, suggesting the wide practical use of SACs in sensing catecholamines-related metabolites.