Real-time Detection Of Hydrogen Peroxide Released From Living Cells By Electrochemical Sensors Based On Fe Nanocomposite Sensing Interface

Hydrogen peroxide(H₂O₂)is an important biomarker related to human metabolism.Real-time detection of H₂O₂ released by living cells is essential for a better understanding of the pathological process of diseases.However,H₂O₂ has the problems of fast diffusion,low concentration,and susceptibility to interference in the cell microenvironment.The advantages of good portability,high sensitivity and good selectivity of electrochemical sensing technology well meet these needs.Since the test object is living cells,the nanomaterials used for testing need to have good cell non-toxicity and biocompatibility.Feelement,as one of the important constituent elements in the human body,has good biocompatibility.Therefore,this article synthesized and designed three nanocomposite materials based on Feelements,constructed an electrochemical biomolecule detection platform with good biocompatibility,and realized quantitative real-time analysis and detection of H₂O₂released by living cells,with the low detection of 57 nM.Further,predict the f M level concentration of H₂O₂ produced by single cells,and discuss its catalytic mechanism in depth.The specific works are as follows:(1)A Cu₂O-mediated AuNPs-FeMOFs was constructed for real-time monitoring of H₂O₂released by living cells.Based on the Cu₂O-MIL-53(Fe)assembled by redox reaction,a stepwise assembly method was used to prepare an electrochemical sensor containing Au^?+(0<?<1)by electrodepositing gold nanoparticles with excellent H₂O₂catalytic reduction activity.The detection range of Au@Cu₂O-MIL-53(Fe)/GCE was 10?M-1520?M,with 1.01?M detection limit and 351.57?A·mM^-1·cm^-2 sensitivity.It had been successfully used to monitor the H₂O₂released by A549 cells in real time under stimulation.Further,the average amount released by a single cells was 0.042 f M by calculated.In addition,the reliability of the biosensor was tested by fluorescently staining the material co-cultured cells and stimulated cells.(2)The preparation process of AuNPs-FeMOFs sensor based on Cu₂O-mediated stepwise assembly was relatively cumbersome,and the conductivity needed to be improved.This chapter used ultrasound to construct a new type of Fe-hemin-MOFs with unique peroxidase activity combined with highly conductive graphene oxide(rGO)reduced by chitosan(CS)for real-time detection of H₂O₂ released by living cells.Based on the strong carboxylate coordination reaction between Fe³⁺and hemin,an electrochemical sensor of Fe-hemin-MOFs/CS-rGO@GCE was constructed,which had the good anti-interference ability to high concentration of 1 mM interferents and low detection limited of 57 nM for H₂O₂ reduction ability.The linear range includes 1-61?M and 61-1311?M,corresponding to the sensitivity of 356.9?A mM^-1cm^-2 and 179.6?A mM^-1cm^-2.The cell growth status of different addition of composite materials in the cell culture process was tested,and the average H₂O₂ released by a single MCF-7 cell under stimulation was monitored in real time as 0.088 pM.(3)To broaden the actual use value of the sensor,an electrochemical sensor with low detection limit,wide detection range and high sensitivity is constructed.KOH-Mxene-regulated carboxylated multi-walled carbon nanotubes with high sensitivity and wide detection range loaded with hemin(MWCNTs/Hemin)nanocatalyst was constructed for real-time detection of live cell release H₂O₂.Utilizing the abundant-OH/O on the surface of KOH-Mxene modified carboxylated MWCNTs/Hemin,a MWCNTs/Hemin/KOH-Mxene nanocomposite with good water dispersibility and electrochemical catalytic performance was obtained.The CV test confirmed that the reaction of the electrode material catalyzes the reduction of H₂O₂was reversible,which controlled by adsorption.And the Laviron equation and Gaussian calculation were used to further calculate the number of electron transfers and pathways for hemin to catalyze the reduction of H₂O₂.The i-t test showed that this electrochemical sensor had a wide detection range of 10-2820?M,a high sensitivity of9.3?A mM^-1cm^-2,and a detection limit of 0.97?M.The composite material was co-cultured with cells to prove its good biocompatibility.The average amount of H₂O₂released by a single MCF-7 live cell under stimulation was 0.017 pM in real-time detection.