Li Qiulan, Li Jitao, Yang Dezhi, Xiang Cheng, Yang Yaling
Faculty of Life Science and Technology, Kunming University of Science and Technology, Kunming 650500, Yunnan, PR China.
Faculty of Life Science and Technology, Kunming University of Science and Technology, Kunming 650500, Yunnan, PR China.
Talanta. 2025 Apr 1;285:127330. doi: 10.1016/j.talanta.2024.127330. Epub 2024 Dec 9.
Endotoxins are found in the outer membrane of all gram-negative bacteria and are a primary cause of endotoxemia, organ failure, and significant harm to human health. Limulus reagent as traditional detection method has certain limitations for the rapid and accurate detection of endotoxin due to the high costs associated with the horseshoe crab used in the sensing process. Herein, a fluorometric and colorimetric dual-mode biosensor was prepared for the rapid and sensitive detection of endotoxin. The chitosan-grafted Fe,Cu-doped carbon dots (CDs) was used to reduce KMnO to produce CS@Fe,Cu/CDs-MnO, which demonstrated distinctive characteristics including high catalytic activity as an artificial nanozyme and a fluorescence quantum yield of 76 %. On the one hand, CS@Fe,Cu/CDs-MnO with the peroxidase-like activity and positively charged property can oxidize 3,3-diaminobenzidine tertrahydrochloride (DAB) and HO to yield a brown product (oxDAB). Additionally, it can interact with negatively charged endotoxin through electrostatic forces, which leads to a reduction in its nanozyme catalytic activity. On the other hand, oxDAB can quench the CS@Fe,Cu/CDs-MnO fluorescence via inner-filter effect and static quenching. Therefore, a dual-mode biosensor that utilizes both fluorescence and colorimetric methods is developed with the aid of a smartphone. It was found that the gray value of absorbance increased linearly with endotoxin concentration in the range of 0.125-175 EU/L with a minimum limit of detection (LOD) of 0.058 EU/L, while the gray value of fluorescence intensity also demonstrated a linear increase upon the addition of endotoxin within the range of 0.05-90 EU/L with a LOD of 0.036 EU/L. The sensor is effectively utilized for detecting endotoxin in real injection samples, achieving acceptable recovery rates between 89.0 % and 101.3 %, with a relative standard deviation (RSD) of no more than 4.51 %. This indicates the reliability of the fabricated biosensor.
内毒素存在于所有革兰氏阴性菌的外膜中，是内毒素血症、器官衰竭以及对人类健康造成重大危害的主要原因。鲎试剂作为传统检测方法，由于传感过程中使用的鲎成本高昂，在内毒素的快速准确检测方面存在一定局限性。在此，制备了一种用于快速灵敏检测内毒素的荧光比色双模式生物传感器。壳聚糖接枝的铁、铜掺杂碳点（CDs）用于还原高锰酸钾以制备CS@Fe,Cu/CDs-MnO，其表现出独特的特性，包括作为人工纳米酶具有高催化活性以及荧光量子产率为76%。一方面，具有类过氧化物酶活性和正电荷性质的CS@Fe,Cu/CDs-MnO可以氧化3,3-二氨基联苯胺四盐酸盐（DAB）和H₂O₂生成棕色产物（oxDAB）。此外，它可以通过静电力与带负电荷的内毒素相互作用，这导致其纳米酶催化活性降低。另一方面，oxDAB可以通过内滤效应和静态猝灭猝灭CS@Fe,Cu/CDs-MnO的荧光。因此，借助智能手机开发了一种同时利用荧光和比色方法的双模式生物传感器。研究发现，吸光度的灰度值在0.125 - 175 EU/L范围内随内毒素浓度呈线性增加，最低检测限（LOD）为0.058 EU/L，而荧光强度的灰度值在0.05 - 90 EU/L范围内加入内毒素后也呈线性增加，LOD为0.036 EU/L。该传感器有效地用于检测实际注射样品中的内毒素，回收率在89.0%至101.3%之间，相对标准偏差（RSD）不超过4.51%。这表明所制备的生物传感器具有可靠性。
