基于电化学阻抗谱的致病菌检测传感器的研究进展.pdf

REVIEWRecent Advances in Electrochemical ImpedanceSpectroscopy-based Pathogenic Bacteria SensingTao Chena,Yuan-Hong Xua,*,Jing-Hong Lib,*aInstitute of Biomedical Engineering,College of Life Sciences,Qingdao University,Qingdao,266071,ChinabDepartment of Chemistry,Key Laboratory of Bioorganic Phosphorus Chemistry&Chemical Biology,Tsinghua University,Beijing,100084,ChinaAbstractPathogenic bacteria have been throwing great threat on human health for thousands of years.Their real-timemonitoring is in urgent need as it could effectively halt the spread of pathogenic bacteria and thus reducing the risk tohuman health.Up till now,diverse technologies such as electrochemistry,optics,piezoelectricity and calorimetry havebeen developed for bacteria sensing.Therein,electrochemical impedance spectroscopy(EIS)-based sensors showgreat potential in point-of-care bacterial analysis because of their low-cost,short read-out time,good reproducibility,and portable equipment construction.In this review,we will primarily summarize the typical applications of elec-trochemical impedance technology in bacteria sensing based on different electrodes in the last three years.As weknow,the electrode materials play an extremely important role in the construction of EIS-based sensors because notonly the immobilization of bio-recognition elements for bacteria,but also the sensitivity,economical efficiency andportability of the as-prepared sensors are mainly determined by the electrode materials.Therefore,in order to providenew researchers a clear preparation process for EIS-based sensors fabricated with different electrodes,we try toclassify the EIS-based sensors according to the different electrode platforms.Moreover,present difficulties,futuredirections and perspectives for their applications are also discussed.It can provide guidance in future study of novelEIS-based sensors for rapid,sensitive and accurate sensing of diverse pathogenic bacteria.Keywords:Electrochemical impedance spectroscopy;Pathogenic bacteria detection;Bio-recognition elements;Elec-trode material1.IntroductionPathogenic bacteria,ubiquitously spreading innearly every corner of human living environment,could lead to serious infectious diseases such asfoodborne disease,urinary system infections aswell as sexually transmitted disease 1e5.There-fore,their accurate,sensitive and rapid detectionhas been a major public health concern.Nowa-days,the developed technologies in laboratory forpathogenic bacteria detection are mainly based ontechnologies including colony morphology recog-nition 6,fluorescence resonance energy transfer7,electrochemistry 8,9,colorimetry 10,nucleicacid-based diagnosis 11,fluorescent and opt-electric technology 12,nanopore technology 13and calorimetry 14.Therein,only three methodsbased on colony,polymerase chain reaction andenzyme linked immunosorbent assay are admittedby Food and Drug Administration,and the devel-oped sensors have been come to commercial ap-plicationsfromlaboratoryprototypes15.Generally,theseapproachesusuallyrelyonmultistep processes including bacterial culturing,isolation and selective enrichment of target or-ganisms to a detectable level.Obviously,thesesensitive but time-consuming(more than 24 h)methods are unsuitable for applications in real-Received 31 October 2022;Received in revised form 5 January 2023;Accepted 17 May 2023Available online 24 May 2023*Corresponding author,Yuan-Hong Xu,Tel:[phone],E-mail address:.*Corresponding author,Jing-Hong Li,Tel:[phone],E-mail address:.https:/doi.org/10.13208/j.electrochem.22180021006-3471/2023 Xiamen University and Chinese Chemical Society.This is an open access article under the CC BY-NC license(http:/creativecommons.org/licenses/by-nc/4.0/).time monitoring and rapid detection of pathogenicbacteria.Therefore,although these methods havecome to real-world implementation,centralizedinstruments and skilled personnel are also essen-tial,limiting their further commercial applicationsin large-scale.To the contrary,electrochemical sensors havealready found great commercial success in thefields of personalized diabetes management 16,because electrochemistry provides the fabricatedsensors with advantages including high specificity,sensitivity,portability and economical efficiency8.Moreover,their fast signal readout and easierminiaturizationmakethem great potential inpoint-of-care bacterial analysis 17.Consequently,electrochemicalimpedancespectroscopy(EIS)-based sensors have been applied for the identifi-cation and determination of bacteria for more than50 years 9,18.The initial utilization of EIS is in thefield of microbiology monitoring such as biofilmformation and growth of overall bacterial 19.Thesystem is generally made of two planar electrodesimmersed in culture medium,which could achievethe real-time detection of bacterial growth densityby monitoring the electrochemical parameters ofthe growth medium 20.Accordingly,dozens ofreal-time commercial products have been devel-oped and the determination of metabolic activity ofbacteria is also realized 21.Based on this mech-anism,EIS-based sensors have also been proved asan effective and informative technique for bacteriadetection,which could reflect the information ofbacterial species and concentrations by monitoringthe occurring reaction on the employed electrodesurface as well as direct tracing the interactionsbetween the bio-receptor and target bacteria 22.In bacteria sensing process,the binding of bac-teria or metabolites produced by bacteria to thesurface of employed electrode could change theelectricalproperties(diffusiveelectrochemicalimpedance,double layer capacitance,and chargetransfer resistance,etc.)of the working electrode,either because of the inherent properties of bac-terial cell membranes or by intercepting diffusingredox-active molecules from interacting with thesurface23.Indetail,iftheelectrochemicalimpedance is obtained in the sensing systemwithoutredoxprobes,themeasuredelectro-chemical impedance signal is a direct reflection ofintact bacteria and could be easily influenced bythe number,morphology and growth stage of thebacteria 9.In the system that contains redoxprobes,the change in faradaic impedance isinstead tested and obtained.Moreover,like gen-eral biosensors,in order to achieve the specificdetection of certain pathogenic bacterium,bio-recognition elements such as antibodies 24e29,aptamers 30e32 and bacteriophages 33e36 arefixed on the surface of different working electrodesas shown in Fig.1 via electrostatic adsorption orcovalent coupling to capture/detectpathogensselectivelyandspecifically.Therefore,theemployed bio-recognition elements endow theobtained sensors with inherent sensitivity anddiscrimination.In the last three years,a series of works on thestudy of EIS-based sensors have been reported anda great process has been made for bacteria sensing.Although several interesting review articles on theuse of EIS in bacterial sensing have been published9,37e39,the rapidly expanding literature reviewin the past three years has not yet been found.Inthis review,we will primarily summarize thetypical applications of EIS in bacteria sensing inthe last three years.Present difficulties,future di-rections and perspectives for the EIS-based sen-sors are also discussed.2.The selection of electrodesThe electrode materials are equally important forthe construction of EIS-based sensors.For suchutilizations,portable electrodes such as glassycarbon electrode,gold electrode,carbon-basedelectrode,screen-printed electrode,indium tinoxide electrode and carbon paste electrode aremost frequently used,which greatly promote thedevelopment of EIS-based sensors.The selectionof electrode material for the construction of EIS-based sensor is usually decided on the cost,com-mercial availability and potential need for surfacemodifications 8.Thus,frequently used materialscontain glassy carbon 30,36,40e42,noble metals43e47 such as gold,silver and platinum,carbon-based materials 24,48e50 such as graphene oxide(GO),carbon nanotubes,carbon nano-walls,andmetal oxides 51,52 such as nickel oxide and in-dium tin oxide,etc.Improvements to electrodeplatforms via modification of materials with largesurface area and conductivity have enabled the as-prepared sensor with enhanced test sensitivity andfast signal readout time.With the miniaturization of electrodes,sophisti-cated interdigitated electrode arrays,which isusually called the second generation of electrode,arereceivinggreatconcerninrecentyears26,29,32,53.The interdigitated electrode arraysare commonly made of two individual electrodestripscontainingmultiplemicroelectrodes8.Therein,each set of microelectrodes could thuswork as a pole for bipolar electrochemical imped-ancetest.ThesensingplatformsbasedonJournal of Electrochemistry,2023,29(6),2218002(2 of 18)interdigitatedelectrodesprovideoutstandingspace efficiency because both spacing and size ofthese electrodes are well-optimized 9.Moreover,the interdigitated electrode arrays have obviousmerits over conventional ones,such as low re-sistances,mall required sensing volumes,quickequilibrations as well as high signal-to-noise ratios54.Herein,we try to classify the EIS-based sen-sors for bacteria sensing according to the electrodeplatforms.As shown in Table 1,the related pa-rameters of the constructed EIS-based sensorsincluding electrode materials,analyte,linear rangeand limit of detection(LOD)are also summarized.2.1.EIS-based sensors for bacterial sensing2.1.1.Glassy carbon electrode(GCE)for EIS-basedsensor developmentIn the fabrication of EIS-based sensors,GCEshave been receiving great concern due to theirsmall thermal expansion coefficient,high chemicalstability,and outstanding air tightness 56,57.Tofurther enhance the conductivity of the workingelectrode and immobilize more bio-recognitionelements on GCE,gold materials such as Aunanoparticles(Au NPs)30,36,40 and gold nano-rods 58 have been applied for GCE modification.Via electrochemical deposition of Au NPs on thesurface of GCE as shown in Fig.2A,Mulchandanisgroup successfully obtained the Au NPs modifiedGCE(GCE-Au)36.Then,M13 bacteriophagewere immobilized on the surface of GCE-Au viacross-linkingreactionof3-mercaptopropionicacidand1-(3-dimethylaminopropyl)-3-ethyl-carbodiimidehydrochloride/N-hydroxysuccini-mide,obtainingaselectivenon-lyticM13bacteriophage-based cytosensor for Escherichia coli(E.coli)sensing.The biosensor achieved a LOD of14CFU/mL.Interestingly,theas-preparedbiosensorexhibitedthesimilarsensitivityinphosphate buffer as in river water samples,indi-cating its good applicability to real samples.Withthe similar methods,anti-protein antibody(IgY)and aptamer were respectively fixed on the surfaceof AuNP-modified GCE,by Roushani et al.40and Dai et al.30.Accordingly,the immuno-sensors were obtained and the successful detectionof S.aureus was achieved.Molecularlyimprintedpolymers(MIPs)areman-made antibodies with customized bindingsites complementary to that of the employedtemplates in both physical and chemical structures59,60.MIPs are found with obvious merits,suchas easy synthesis,economic efficiency,and long-time chemical and physical stabilities in com-paration with the natural antibodies.Thus,MIPshave been applied for the construction of varioussensing platforms in diverse molecules sensing61e63.As shown in Fig.2B,the molecularlyimprintedsensorshaveachievedthespecificdetection of bacteria with the help of MIPs.Recently,combining with MIPs and EIS,Biansgroup reported the preparation of a reusablesensor for rapid determination of pathogenic bac-teria based on bacteria-imprinted polythiophenefilm(BIF)employingStaphylococcusaureus(S.aureus)as an imprinting template 41.The BIF,as apolymer layer for specific recognition of S.aureus,is deposited on the surface of a GCE via electro-copolymerization of TE monomer in the presenceof S.aureus(template)and followed by templateremoval.When the S.aureus rebinds on the BIF,the electrochemical impedance of the workingelectrode is increased.Accordingly,the BIF-basedFig.1.The general fabrication process of EIS-based bacterial sensors via the modification of different electrodes(glassy carbon,Au,carbon,screen-printed,indium tin oxide,and interdigitated electrode)using different recognition elements(antibody,mannose,MIPs,bacteriophage,lectin,aptamer).Journal of Electrochemistry,2023,29(6),2218002(3 of 18)Table 1.Parameters of the constructed sensors including electrode material,analyte,linear range and LOD in the last three years.Ref.LODLinear rangeAnalyteElectrodeGCEE.coli103e107500 CFU/mLCFU/mL56Gold modified GCEE.coli10e10514 CFU/mLCFU/mL36AuNP modified GCES.aureus10e1073.3 CFU/mLCFU/mL40Dual-aptamer-based sand-wich GCES.aureus1?101to 1?1052 CFU/mLCFU/mL30Gold nanorods modified GCES.aureus1.8?103to 1.8?107CFU/mL2.4?102CFU mL583-Thiopheneethanol modifiedGCES.aureus10e1074 CFU/mLCFU/mL41Synthetic receptor-trans-ducing platform couplingE.coli120 CFU/mL1000 CFU/mL55GCEAcinetobacter baumannii10?1to 1040.030 CFU/mLCFU/mL42Gold electrodesE.coli;L.innocua;S.aureus;S.Typhimurium1.5 to 1.5?103;1.5to 1.5?104;1.5 to 1.5?105;15 to 1.5?104CFU/mL1.5;1.5;1.5;15 CFU/mL45Gold electrodeS.typhimurium and E.coli6.859N/A?1023L$mole1and2.054?1017L$mole143Screen-printed gold electrodeSalmonella15 to 2.57spp.?1075 CFU/mLCFU/mL463D gold nano-/microislandsand graphene electrodesE.coli,P.putida,and S.epidermidis2?10 to 2?105,2?10 to2?104,and 1?102to1?105CFU/mL20 CFU/mL44Gold disk electrodeSalmonella2?10 to 2?106/2?102to2?10517 CFU/mL;1.3?102CFU/mL;1 CFU/mL33Lipid membrane modifiedgold electrodeE.coli10DNA?9to 10?19mol$L?110?19mol$L?165Ag electrodesE.coliandPseudomonasaeruginosa500Up to 500 CFU/mLe1000 CFU/mL47Graphenic carbon electrodesE.coli and S.aureus2e2 CFU/mL20 CFU/mL50Reduced graphene oxide-carbon electrodeS.mutans,A.viscosus,and L.fermentumN/AN/A48Boron-doped carbon nano-walls electrodesPseudomonas syringae pv.lachrymans3.25?100to3.25?108CFU/mL119 CFU/mL24Carbon nanotube-basedelectrodeS.aureus102e1071.23CFU/mL?102CFU/mL;1.29?102CFU/mL34SPEE.coli102e10810 CFU/mLCFU/mL66SPEE.coli1DNA?10?10mmol$L?1to1?10?5mmol$L?11.95?10?15mmol$L?170Screen-printed carbonelectrodesS.aureus10e1083 CFU/mLCFU/mL31Screen printed goldelectrodesS.aureus10?101to 10?107CFU/mL101.58CFU/mL68GlycoMXene screen printedelectrodesE.coli101e10810 CFU/mLCFU/mL67ITO coated polyethyleneterephthalate(ITO:PET)Pseudomonas aeruginosaN/AN/A51Fluorine doped tin oxideelectrodeSalmonella gallinarum,andSalmonella pullorum(1e1?105cells)with 37and 25 viable cells51 and 37 cells,respectively infaecal samples and218 and 173 cells,respectively inmeat samples.52Anti-E.coliO157:H7 antibody-modified CPEE.coli1?10?1to 1?106CFU/mL0.1 CFU/mL81Polyaniline nanofibers modi-fied filter paper substrateS.aureus,E.coli,P.aeruginosaN/AN/A35(continued on next page)Journal of Electrochemistry,2023,29(6),2218002(4 of 18)sensor could determinate S.aureus in the range of10e107CFU/mL witha LOD of 4 CFU/mL.Impressively,the as-prepared sensing platfor