Robust Nanozyme-Enzyme Nanosheets-Based Lactate Biosensor for Diagnosing Bacterial Infection in Olive Flounder (Paralichthys olivaceus)

Abstract

Bacterial infections in fish farms increase mass mortality and rapid detection of infection can help prevent its widespread. Lactate is an important biomarker for early diagnosis of bacterial infections in farmed olive flounder (Paralichthys olivaceus). To determine the lactate levels, we designed a disposable amperometric biosensor based on Prussian blue nanozyme and lactate oxidase (LOX) entrapped in copolymer-reduced graphene oxide (P-rGO) on screen-printed carbon electrodes. Because LOX is inherently unstable, P-rGO nanosheets were utilized as a base matrix to immobilize it. After optimization in terms of enzyme loading, operating potential, and pH, the biosensor displayed maximum current responses within 5 s at the applied potential of –0.1 V vs. internal Ag/AgCl. The biosensor had Langmuir-type response in the lactate concentration range from 10 µM to 1.6 mM, a dynamic linear response range of 10–100 µM, a sensitivity of 15.9 µA mM−1 cm−2, and a lower detection limit of 3.1 µM (S/N = 3). Additionally, the biosensor featured high reproducibility, good selectivity, and stability till four weeks. Its practical applicability was tested in olive flounder infected by Streptococcus parauberis against the uninfected control. The results were satisfactory compared to those of a standard colorimetric assay kit, validating our method.

A Study on the Mechanism and Properties of a Self-Powered H2O2 Electrochemical Sensor Based on a Fuel Cell Configuration with FePc and Graphene Cathode Catalyst Materials

“…Hydrogen peroxide (H 2 O 2 ) is not only widely used in the medical, health, industrial, and domestic fields, but it also plays an important role in the regulation of biological activities [ 1 , 2 , 3 , 4 ]. Moreover, H 2 O 2 is a product of oxidoreductase-catalyzed transformations; its quantification is used for the detection of other biologically important markers, e.g., glucose, lactate, uric acid, and glutamate [ 5 , 6 , 7 ]. Therefore, the detection of H 2 O 2 has been widely studied with different methods, such as fluorescence, spectrophotometry, and electrochemistry [ 8 , 9 , 10 ].…”

“…The electrodes are therefore modified with the catalyst to improve the thermodynamics and kinetics of the electrode reactions as well as the analytical characteristics of the respective sensors, such as natural enzymes [ 3 , 8 , 19 , 20 ], non-enzymatic electrocatalysts including metal nanostructures [ 21 , 22 , 23 ], carbonaceous nanomaterials, enzyme mimetics, and nanozymes such as metallocomplexes and Prussian blue [ 6 , 7 , 8 , 24 , 25 , 26 , 27 ]. Of the aforementioned materials, enzymatic catalysts have advantages in terms of specificity and high catalytic activity, which are determined by the properties of the enzymes.…”

The Nanozyme Revolution: Enhancing the Performance of Medical Biosensing Platforms

“…Similar cascade reactions have also been reported for the detection of other oxidase substrates such as acetylcholine, lactate, xanthine, uric acid, and other relevant biomolecules by coupling their corresponding oxidase enzyme with a peroxidase nanozyme. [59][60][61][62] In recent years, the application of nanozymes has been extended to more complex bioassays such as immunosorbent assays, electrochemical biosensors and even in vivo sensing platforms and each of these are discussed in turn below…”