组成图示
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传感器类型
电化学生物传感器
检测对象
乙醇（ethanol，酒精，BAC）；样品基质：全血/血液样品（whole blood, blood samples）
检测原理
该传感器基于ADH催化乙醇氧化：乙醇在ADH和NAD+存在下被氧化为乙醛，同时NAD+被还原为NADH。NADH在MB介导下于工作电极发生低过电位氧化，释放电子；电子经MB、GNPs、MWCNT和Nafion复合膜传递至SPE碳工作电极，在-0.13 V（vs. Ag/AgCl）下以计时电流法读取安培电流。乙醇浓度越高，生成的NADH越多，稳态电流越大。GNPs和MWCNT通过高导电性、大电活性面积和协同电子传递放大电流，MB降低NADH氧化过电位并减少干扰，亲水膜限制样品体积，实现小体积全血BAC检测。
检测灵敏度
LOD: 5.0 × 10^-5 mol/L (S/N = 3)；线性范围: 2.0 × 10^-4 to 25 × 10^-3 mol/L；标准曲线: Δi (mA) = 0.09347 + 0.12141C (mmol/L)；r = 0.99498
效应效果
在20 mmol/L干扰物测试中，葡萄糖、甲醇、乳酸、丙酮、抗坏血酸、正丙醇、异丁醇、正丁醇和异丙醇的干扰率分别为3.31%、6.41%、4.52%、-4.92%、-4.35%、5.82%、4.97%、5.16%和5.63%，抗干扰性良好。4 ℃保存20天和30天后响应电流分别下降3.6%和5.8%；批内CV为4.1%，批间CV为4.7%。响应时间小于15 s，表观Km为5.3 mmol/L。对60例酒驾血液样品（50例活体、10例死亡）检测，与气相色谱法线性相关方程为Y=1.03986X-0.2644，r=0.99621，表明可用于法医BAC快速筛查。
传感器的构成
- 基底/换能器电极：PVC基底上的丝网印刷电极（SPE），含碳工作电极、碳对电极和Ag/AgCl参比电极，提供三电极电化学换能。
- 纳米复合修饰层：GNPs–MWCNT–Nafion-117纳米复合膜，滴涂于工作电极，GNPs与MWCNT协同导电并增大电活性面积，Nafion成膜并吸附介体。
- 电子介体层：Meldola's blue（MB），通过循环伏安吸附于GNPs–MWCNT–Nafion膜，介导NADH在低过电位氧化并传递电子。
- 识别/催化元件层：乙醇脱氢酶（ADH）与烟酰胺腺嘌呤二核苷酸（NAD+）混合酶液，滴涂于MB修饰膜，ADH催化乙醇氧化并生成NADH。
- 封闭/稳定剂：牛血清白蛋白（BSA），与ADH和NAD+共固定，稳定酶并减少非特异吸附。
- 样品反应腔：中心带孔的亲水膜覆盖工作区，形成5 mL反应池，限制样品体积并允许血液接触电极。
- 信号读出：mAutolab III电化学工作站，在-0.13 V（vs. Ag/AgCl）下采用计时电流法读取安培电流。
中文摘要
本研究构建了一种用于小体积样品检测血醇浓度（BAC）的新型可抛弃安培生物传感器试纸条。该传感器以丝网印刷电极（SPE）为基底，先在工作电极上修饰由Nafion-117、多壁碳纳米管（MWCNT）和金纳米粒子（GNPs）组成的GNPs–MWCNT–Nafion纳米复合膜；随后以Meldola's blue（MB）作为电子传递介体，并将乙醇脱氢酶（ADH）与烟酰胺腺嘌呤二核苷酸（NAD+）依次固定于复合膜上；最后在工作区外覆盖中心带孔的亲水膜，形成5 mL反应池，得到亲水膜/ADH–NAD+/GNPs–MWCNT–MB–Nafion/SPE试纸条。通过吸液方式精确取5 mL血液样品即可完成BAC检测。实验优化了工作电位、pH、温度及干扰物等条件。结果表明，该传感器准确度和稳定性良好，线性范围为2.0×10^-4至25×10^-3 mol/L，检出限为5.0×10^-5 mol/L（S/N=3）。在真实血液样品检测中，其结果与气相色谱法具有良好相关性，可用于BAC分析。
英文摘要
A novel disposable amperometric biosensor strip for determination of blood alcohol concentration (BAC) with small volume of sample has been constructed using screen-printed electrodes (SPE), nanocomposite film and alcohol dehydrogenase (ADH). Firstly, the GNPs-MWCNT-Nafion nanocomposite film modified on a working electrode was made of Nafion-117, multi-wall carbon nanotubes (MWCNT) and gold nanoparticles (GNPs). After that Meldola's blue (MB) acted as an electron transfer mediator and the mixed solution of ADH, nicotinamide adenine dinucleotide (NAD(+)) were modified in order on the nanocomposite film. At last, a hydrophilic membrane which had an eyehole in center was placed at the outermost of the working area to make a reaction tank of 5 μL, then the hydrophilic membrane/ADH-NAD(+)/GNPs-MWCNT-MB-Nafion/SPE was prepared. The detection of BAC can be accomplished with 5 μL of blood sample obtained precisely by siphonage. Optimum conditions of the biosensor were experimentally determined by varying several important parameters: working potential, solution pH value, environmental temperature and interferences. Experimental results indicated that the biosensor possessed a good accuracy and stability, the linear response range was 2.0 × 10(-4) to 25 × 10(-3)mol/L and the detection limit of the biosensor was 5.0 × 10(-5)mol/L (S/N=3). In the measurement of blood samples, the proposed biosensor had excellent detection performance for measuring BAC and showed a good correlation with gas chromatography. The prepared biosensor strip can be valid for the analysis of BAC.