Dual-mode colorimetric-fluorescence biosensor for endotoxin detection based on CS@Fe,Cu/CDs-MnO nanomaterials.

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.