Monitoring copper (Cu) species in bio-samples is critical for the investigation and auxiliary diagnosis of Cu-metabolism disorders. Herein, a target-triggered DNAzyme walker, consisting of Carboxyfluorescein (FAM)-labeled substrates of the Cu-DNAzyme (Cu-Sub) coated gold nanoparticles (AuNPs) (AuNPs@FAM-Cu-Sub track) and the enzyme chain of the Cu-DNAzyme (Cu-Enzy) functionalized magnetic beads (MBs@Cu-Enzy 3D walking unit), was designed for the sensitive detection of copper species. Upon addition of copper ion (Cu²⁺), DNAzyme was activated and triggered the walking process, thereby releasing FAM-labeled fragments and recovering fluorescence. When compared with free-running Cu-Enzy or co-conjugated Cu-Enzy on AuNPs walking units, the MBs@Cu-Enzy walking unit exhibited a higher cleavage efficiency, shorter detection time (40 min), and a lower limit of detection (LOD = 3 nM). This improved performance was mainly attributed to multivalent interactions between the 3D track and the 3D walking unit which dramatically increased the affinity and cleavage efficiency. Importantly, good accuracy was observed for copper species analysis in clinical serum samples (n = 130), relative to other instrumentation, inductively coupled plasma atomic emission spectrometry (ICP-AES) (P > 0.01). The serum detection data indicated that total, exchangeable and relative exchangeable Cu variables could serve as potential indicators for the auxiliary diagnosis of Cu-related disorders. Thus, our target-triggered DNAzyme walker provides improved technical support for the in-depth study of clinical Cu metabolic-related diseases.

监测生物样品中的铜（Cu）物种对于研究和辅助诊断Cu代谢紊乱至关重要。在这里，一个目标触发的DNAzyme步行器，由羧基荧光素（FAM）标记的Cu-DNAzyme（Cu-Sub）包被的金纳米颗粒（AuNPs）（AuNPs @ FAM-Cu-Sub轨道）的底物和酶链组成。 Cu-DNAzyme（Cu-酶）功能化的磁珠（MBs @ Cu-Enzy 3D步行单元）设计用于灵敏地检测铜物种。添加铜离子（Cu ²⁺），DNAzyme被激活并触发步行过程，从而释放FAM标记的片段并恢复荧光。与在AuNPs行走单元上自由运行的Cu-Enzy或共轭Cu-Enzy相比，MBs @ Cu-Enzy行走单元显示出更高的裂解效率，更短的检测时间（40分钟）和更低的检测限（ LOD = 3 nM）。这种改进的性能主要归因于3D轨道和3D步行单元之间的多价相互作用，从而大大提高了亲和力和切割效率。重要的是，与其他仪器，电感耦合等离子体原子发射光谱法（ICP-AES）相比，在临床血清样品（n = 130）中铜的种类分析观察到了良好的准确性（P> 0.01）。血清检测数据表明，总的，可交换的和相对可交换的铜变量可以作为辅助诊断铜相关疾病的潜在指标。因此，我们的目标触发的DNAzyme沃克为深入研究临床与铜代谢相关疾病提供了更好的技术支持。