Direct real-time measurements of superoxide release from skeletal muscles in rat limbs and human blood platelets using an implantable Cytochrome C microbiosensor
Direct real-time measurements of superoxide release from skeletal muscles in rat limbs and human blood platelets using an implantable Cytochrome C microbiosensor
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DOI：
10.1016/j.bios.2023.115664
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发表时间：
2023-09-07
影响因子：
12.6
通讯作者：
Andreescu, Silvana
中科院分区：
文献类型：
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作者：
Deshpande, Aaditya S.;Muraoka, Wayne;Andreescu, Silvana
氧化应激和超氧阴离子(O-2(中心点))的过度积累是许多病理情况和几种疾病发生的原因。实时监测(O-2(中心点))的释放对于评估在这些条件下的氧化应激程度是重要的。在这里，我们介绍了一种坚固的(O-2(中心点))生物传感器的设计、制造和表征，方法是在细胞色素C(Cytc)偶联的金丝电极上沉积一层均匀的L-半胱氨酸。用导电原子力显微镜(c-AFM)研究了固定化层，发现在金表面有稳定且均匀分布的氧化还原蛋白，显示为导电性和表面地形图。该生物传感器可在0.15V的外加电压下检测O-2(中心点-)，灵敏度为42.4 nA/mU M，检测下限为2.4 nM。将该生物传感器应用于脑缺血再灌注损伤(IRI)后激活的人血小板和骨骼肌中O-2(中心点)的实时释放，证实了该生物传感器在复杂生物系统中的稳定性和稳健性。结果表明，这些生物传感器能够实时监测(O-2(中心点))的释放，并在容易转化为人类病理的模型中估计氧化损伤的程度。
Oxidative stress and excessive accumulation of the superoxide (O-2(center dot-)) anion are at the genesis of many pathological conditions and the onset of several diseases. The real time monitoring of (O-2(center dot-)) release is important to assess the extent of oxidative stress in these conditions. Herein, we present the design, fabrication and characterization of a robust (O-2(center dot-)) biosensor using a simple and straightforward procedure involving deposition of a uniform layer of LCysteine on a gold wire electrode to which Cytochrome C (Cyt c) was conjugated. The immobilized layers, studied using conductive Atomic Force Microscopy (c-AFM) revealed a stable and uniformly distributed redox protein on the gold surface, visualized as conductivity and surface topographical plots. The biosensor enabled detection of (O-2(center dot-)) at an applied potential of 0.15 V with a sensitivity of 42.4 nA/mu M and a detection limit of 2.4 nM. Utility of the biosensor was demonstrated in measurements of real time (O-2(center dot-)) release in activated human blood platelets and skeletal rat limb muscles following ischemia reperfusion injury (IRI), confirming the biosensor's stability and robustness for measurements in complex biological systems. The results demonstrate the ability of these biosensors to monitor real time release of (O-2(center dot-)) and estimate the extent of oxidative injury in models that could easily be translated to human pathologies.