Reply to the Correspondence on “Carbon-Dot-Based Dual-Emission Nanohybrid Produces a Ratiometric Fluorescent Sensor for in Vivo Imaging of Cellular Copper Ions”

The Correspondence by Gharib et al. challenges our 2012 study on CQD-based Cu²⁺ sensors but introduces no new experimental data. Their failure to reproduce our results stems directly from fundamental errors in materials synthesis. Specifically, they failed to properly synthesize either key material. Regarding AE-TPEA, the core innovation of our work, they deviated from our published protocol, failed to synthesize and validate the identity and purity of the product, and did not report the deviation from the protocol in the Correspondence. Without authentic AE-TPEA, the subsequent experiments are scientifically meaningless. Regarding CQDs, we followed the established method of Li et al., a protocol independently reproduced by numerous laboratories worldwide. The authors failed to synthesize the proper fluorescent CQDs because they did not strictly follow the experimental conditions, not because of a flaw in our approach. Furthermore, their own data are internally inconsistent. Three parallel syntheses yielded conflicting results across UV-vis, fluorescence, FTIR, TEM, DLS, and zeta potential measurements, exposing a lack of experimental control. Finally, our work has been widely validated. Over a decade of independent applications, many research groups have confirmed the reproducibility and utility of our approach.