【摘要】 目的 开发一种基于分子印迹技术（molecularly imprintedpolymers, MIPs）的高灵敏度、高选择性电化学探针，用于25-羟基维生素D₃[25-（OH）D₃]的精准检测，以满足维生素D代谢相关疾病的诊断需求。方法 通过逐层修饰丝网印刷电极（screen-printed electrode, SPE），构建Co₃O₄@还原氧化石墨烯（reducedgrapheneoxide,rGO）@Pt@Au复合基底，以增强导电性。结合聚吡咯的电沉积与模板分子洗脱，成功形成了具有特异性识别能力的印迹空腔。通过优化扫描速率、沉积循环次数、孵育时间和温度，平衡了膜层结构、电子传递效率与特异性结合能力。采用方波伏安法（square wave voltammetry,SWV）对该传感器的响应性能进行评估，分析其灵敏度、选择性及重现性。结果 扫描电子显微镜（scanning electronmicroscope,SEM）表征显示，模板洗脱后电极表面形成清晰的印迹空腔，该空腔可在100nmol/L至1mmol/L范围内进行靶标检测，其检测限为26.76 nmol/L。该检测策略对10倍浓度干扰物无特异性响应，且3批次电极重现性相对标准偏差为3.2%。加标实验也显示该检测策略具有较好的实际应用能力。结论 本研究成功构建了一种灵敏度高、抗干扰性强、重现性优异的新方法，为临床上25-（OH）D₃的检测提供了一种新思路。更多还原

【Abstract】 Objective To develop a highly sensitive and selective electrochemical probe based on molecular imprinting polymers（MIPs） technology for the precise detection of 25-hydroxyvitamin D₃ [25-（OH）D₃], aiming to meet the diagnostic needs of vitamin D metabolism-related diseases. Methods The Co₃O₄@reduced graphene oxide（rGO）@Pt@Au composite substrate was constructed by layer-by-layer modification of screen-printed electrode（SPE） to enhance conductivity. Molecularly imprinted cavities were successfully formed through electropolymerization of polypyrrole and template molecule elution, imparting specific recognition capabilities. By optimizing the scan rate, deposition cycles, incubation time, and temperature, the balance between membrane structure, electron transfer efficiency, and specific binding ability was achieved. The sensor’s response performance was evaluated using square wave voltammetry（SWV）, and its sensitivity, selectivity, and reproducibility were analyzed.Results Scanning electron microscopy（SEM） characterization revealed that after template elution, clear imprinted cavities were formed on the electrode surface, which could detect the target in the range of 100 nmol/L to 1 mmol/L, with a detection limit of 26.76 nmol/L. This detection strategy showed no specific response to interference at 10 times the concentration, and the reproducibility of the electrodes across three batches, with a relative standard deviation of 3.2%, indicated the reliability of the method. The spiked experiments also show its good practical application ability. Conclusion This study successfully established a new method with high sensitivity, strong anti-interference ability, and excellent reproducibility, providing a novel approach for the clinical detection of 25-（OH）D₃.更多还原