## 3 Results and Discussion
### Calibration Curves for Adr in Tris Buffer  
After confirming that the shifts observed in the CVs due to individual components of the Tris buffer matrix were negligible, we proceeded to construct calibration curves for Adr in the complete Tris buffer system. This step was critical for evaluating the performance of our electrochemical sensor in detecting Adr under conditions that mimic the ionic and chemical complexity of artificial cerebrospinal fluid. To construct the calibration curves, we systematically varied the concentration of Adr in the Tris buffer and recorded CVs for Adr transfer using both ionophores, DB24C8 and DB18C6. The CVs were analyzed to determine the relationship between Adr concentration and the observed electrochemical signal. Interestingly, although there was no significant increase in the current with increasing Adr concentration, a clear and reproducible increase was observed in the shifts between the half-wave potential for Adr and the Tris buffer as the concentration of Adr increased (Figure 5). With DB24C8, the shift in E\({}_{1/2}\) exhibited a well-defined trend, increasing with Adr concentration and eventually plateauing, resembling a classical calibration curve. This behavior highlights the high specificity and sensitivity of DB24C8 toward Adr. A similar trend was observed with DB18C6 (Figure S2), although the regression value of the calibration curve was slightly lower compared to DB24C8. Despite this difference, both ionophores demonstrated their capability to detect Adr reliably. Additionally, the log-transformed data revealed an excellent linear relationship between the potential shift and the Adr concentration (Figure 5b). For DB24C8, the calibration curve yielded an R\({}^{2}\) value of 0.98, with a calculated limit of detection (LOD) of 5 pM. Similarly, DB18C6 demonstrated an R\({}^{2}\) value of 0.95, with an LOD of 50 pM (Figure S2). These results indicate that our sensor achieves high sensitivity and precision in detecting Adr, even in the complex Tris buffer matrix. Notably, the LODs reported here are well within the physiologically relevant concentration range of Adr, making these ionophores highly suitable for neurochemical applications. Furthermore, while DB24C8 and DB18C6 are known to facilitate the transfer of other cations such as potassium and calcium, our findings confirm that these transfers do not interfere  
Figure 5: (a) Calibration plot illustrating the relationship between the peak shift and the concentration of Adr (5 pM to 500 \(\upmu\)M) in Tris buffer in the presence of DB24C8. (b) Log–log plot of peak shift versus Adr concentration, showing a power-law relationship. This highlights the dynamic range and scalability of the nanopject sensor. Together, these plots validate the robustness and accuracy of the electrochemical approach for Adr quantification in Tris buffer. with Adr detection. This selectivity is a crucial advantage of our approach, ensuring accurate Adr measurement in matrices with competing ions. The importance of this calibration approach lies in its ability to provide reliable Adr detection in a highly complex environment. Unlike conventional methods, which often rely on direct current measurements, our technique leverages shifts in E\({}_{1/2}\) to quantify the Adr concentration. This novel method not only enhances sensitivity but also offers robustness against potential interference in complicated biological matrices. The successful calibration of our sensor in Tris buffer sets the stage for further testing in real biological samples, ultimately advancing its applicability for in vivo and in vitro neurochemical studies. While our approach is less intricate than redox-based methods, it is best suited for charged molecules. Extending this technique to neutral analytes would require additional mediators or sensor modifications to facilitate their detection. Furthermore, when used with real biological samples such as blood or urine, the interface may become blocked by matrix components, potentially compromising stability over time.