传感器类型
电化学生物传感器
检测对象
葡萄糖(glucose);样品基质:磷酸盐缓冲液(PBS)及10倍稀释人血清
检测原理
葡萄糖在GOx催化下与O2反应生成H2O2;H2O2进入PNT包埋的HRP,将HRP还原态氧化为氧化态,氧化态HRP再催化对苯二酚(HQ)氧化为苯醌(BQ),BQ在电极表面接受电子和质子还原为HQ,形成循环电子传递。该过程在0.03 V低电位下产生阴极电流,电流随葡萄糖浓度增加而增大。PNT提供生物相容纳米环境并可能促进电子转移,AuNP增大有效表面积,双酶级联与介质循环实现信号放大。
检测灵敏度
LOD: 73.1 mM;线性范围: 0.5–2.4 mM;灵敏度: 0.3 mA M^-1;R^2 = 0.994
效应效果
传感器对0.5 mM抗坏血酸、蔗糖、半乳糖和乳糖的响应可忽略,低工作电位0.03 V与Nafion层提高选择性。同一电极8次校准斜率RSD为1.95%,三个电极RSD为5.74%;4°C PBS中保存一个月保留85%初始响应,响应时间约9–10 s。人血清10倍稀释后检测结果与YSI 2300 STAT plus分析仪一致,相对偏差为0.54%、3.88%和7.69%。其灵敏度0.3 mA M^-1优于部分文献报道,适用于糖尿病血糖检测。
传感器的构成
- 基底/换能器电极:多晶金电极(Au electrode,3 mm),经氧化铝抛光、超声清洗和piranha处理,作为工作电极与电子转导基底。
- 纳米材料修饰层:金纳米粒子(AuNP),在0.1 M KNO3含3 mM HAuCl4中于-200 mV电沉积330 s,增大电极表面积并增强固定。
- 自组装单分子层:3-巯基丙酸(MPA)与1-十四烷硫醇(TDT)二元SAM(MPA:TDT=9:1,总1 mM),提供羧基锚定位点和疏水微环境。
- 交联活化层:EDC/NHS(1 mM EDC、3 mM NHS)活化MPA末端羧基,与PNT氨基形成酰胺键。
- 识别/催化元件:葡萄糖氧化酶(GOx,3 mg/mL)与肽纳米管包埋辣根过氧化物酶(PNT/HRP)共固定,GOx氧化葡萄糖产生H2O2,HRP催化H2O2氧化介质。
- 信号标记/电子介质:对苯二酚(HQ)作为电子介质,在HRP氧化态还原循环中传递电子。
- 封闭/抗干扰层:Nafion(0.5 wt%)覆盖电极表面,减少干扰物并稳定酶。
中文摘要
本研究开发了一种新型安培葡萄糖生物传感器,采用仿生肽纳米管(PNT)作为酶的包埋模板。辣根过氧化物酶(HRP)被包埋于PNT中,葡萄糖氧化酶(GOx)与PNT共同固定于金纳米粒子(AuNP)修饰电极上。电极表面形成由3-巯基丙酸(MPA)和1-十四烷硫醇(TDT)组成的二元自组装单分子层(SAM),用于固定PNT和GOx。所得电极可为酶提供生物相容的纳米环境,在较长时间内维持增强的酶活性,并可能通过PNT促进酶与电极之间的直接电子转移。在实验室条件下,从检测限、灵敏度、pH、响应时间、选择性、重现性和稳定性等方面评价了传感器性能,并进行了实际样品检测。AuNP-SAM-PNT/HRP-GOx复合电极对葡萄糖在0.5–2.4 mM范围内呈线性响应,R^2为0.994;最大灵敏度为0.3 mA M^-1,重现性(RSD)为1.95%。PNT包埋酶在储存一个月后仍保留85%的初始电流响应。
英文摘要
A novel amperometric glucose biosensor was developed using the bio-inspired peptide nanotube (PNT) as an encapsulation template for enzymes. Horseradish peroxidase (HRP) was encapsulated by the PNT and glucose oxidase (GO(x)) was co-immobilized with the PNT on a gold nanoparticle (AuNP)-modified electrode. A binary SAM of 3-mercaptopropionic acid (MPA) and 1-tetradecanethiol (TDT) was formed on the surface of the electrode to immobilize the PNT and GO(x). The resulting electrode appeared to provide the enzymes with a biocompatible nanoenvironment as it sustained the enhanced enzyme activity for an extended time and promoted possible direct electron transfer through the PNT to the electrode. Performance of the biosensor was evaluated in terms of its detection limit, sensitivity, pH, response time, selectivity, reproducibility, and stability in a lab setting. In addition the sensor was tested for real samples. The composite of AuNP-SAM-PNT/HRP-GO(x) to fabricate a sensor electrode in this study exhibited a linear response with glucose in the concentration range of 0.5-2.4mM with a R(2)-value of 0.994. A maximum sensitivity of 0.3 mA M(-1)and reproducibility (RSD) of 1.95% were demonstrated. The PNT-encapsulated enzyme showed its retention of >85% of the initial current response after one month of storage.