Ito Kentaro, Inoue Kumi Y, Ito-Sasaki Takahiro, Ikegawa Miho, Takano Shinichiro, Ino Kosuke, Shiku Hitoshi
Department of Frontier Science for Advanced Environment, Graduate School of Environmental Studies, Tohoku University, 6-6-11-604 Aramaki Aoba, Sendai 980-8579, Japan.
Center for Basic Education, Faculty of Engineering, University of Yamanashi, 4-3-11 Takeda, Kofu 400-8511, Japan.
Micromachines (Basel). 2023 Jan 27;14(2):327. doi: 10.3390/mi14020327.
The amebocyte lysate (LAL) reaction-based assay, the most commonly used endotoxin detection method, requires a skilled technician. In this study, to develop an easy-to-use and highly sensitive endotoxin sensor, we created an electrochemical endotoxin sensor by using an interdigitated array electrode (IDAE) device with advantages of amplifiable signals via redox cycling and portability. We added Boc-Leu-Gly-Arg--aminophenol (LGR-pAP) as an electrochemical substrate for an LAL reaction and detected -aminophenol (pAP) released from LGR-pAP as a product of an endotoxin-induced LAL reaction via an IDAE device. The IDAE device showed a great redox cycling efficiency of 79.8%, and a 4.79-fold signal amplification rate. Then, we confirmed that pAP was detectable in the presence of LGR-pAP through chronoamperometry with the potential of the anode stepped from -0.3 to 0.5 V vs. Ag/AgCl while the cathode was biased at -0.3 V vs. Ag/AgCl. Then, we performed an endotoxin assay by using the IDAE device. Our endotoxin sensor detected as low as 0.7 and 1.0 endotoxin unit/L after the LAL reaction for 1 h and 45 min, respectively, and these data were within the cut-off value for ultrapure dialysis fluid. Therefore, our highly sensitive endotoxin sensor is useful for ensuring medical safety.
基于鲎试剂（LAL）反应的检测方法是最常用的内毒素检测方法，需要技术熟练的技术人员。在本研究中，为开发一种易于使用且高度灵敏的内毒素传感器，我们利用具有通过氧化还原循环放大信号和便携性优点的叉指阵列电极（IDAE）装置创建了一种电化学内毒素传感器。我们添加了Boc-Leu-Gly-Arg-对氨基苯酚（LGR-pAP）作为LAL反应的电化学底物，并通过IDAE装置检测从LGR-pAP释放的对氨基苯酚（pAP），其作为内毒素诱导的LAL反应的产物。IDAE装置显示出79.8%的出色氧化还原循环效率和4.79倍的信号放大率。然后，我们通过计时电流法证实，在阳极电位相对于Ag/AgCl从-0.3 V步进到0.5 V而阴极相对于Ag/AgCl偏置在-0.3 V的情况下，在LGR-pAP存在下可检测到pAP。然后，我们使用IDAE装置进行了内毒素检测。我们的内毒素传感器在LAL反应1小时和45分钟后分别检测到低至0.7和1.0内毒素单位/升，这些数据在超纯透析液的临界值范围内。因此，我们的高灵敏度内毒素传感器对于确保医疗安全很有用。
