Research On Electrochemical Biosensors Based On Screen Printed Electrodes

The research on biomacromolecules contributes to understand the mechanism of the life activities and acquire the biological and chemical information in the process of life.It is also of great significance to study biomolecules in the clinical diagnosis and treatment of diseases.Currently,how to detect these biomolecules sensitively,quickly and efficiently is a very important problem in the life science field.Electrochemical biosensors combine the sensitivity of electrochemical transducers with the high specificity of biological recognition processes,thus they have the advantages of fast response,high sensitivity,good selectivity,etc.Also,they are usually cheap,low cost,and well adapted to miniaturization,and therefore electrochemical biosensors can provide an attractive means for the above problem.Bioactive materials,such as enzyme,nucleic acid,antigen and antibody,cell etc.,are fixed on the surface of electrode.Such substances can specifically recognize their analytes,at the same time they don't impede the free diffusion of analytes.The reaction of bioactive substances and the analytes produces a product in the form of a biological or chemical substance,and a transducer changes the product of reaction into usable data.The general performance and usefulness of electrochemical sensors are often dictated by the various electrochemical transduction mechanisms and the surface immobilization techniques.Electrochemical sensors use a variety of electrodes as a transducer.Since reactions are generally detected only in close proximity to the electrode surface,the electrodes themselves play a crucial role in the performance of electrochemical biosensors.Based on the chosen function of a specific electrode,the electrode materials,its surface modification or its dimensions greatly influence the detection ability.No matter which immobilization techniques,the activity of bioactive molecules and the stability of the biosensor must be maintained.Therefore,an electrode substrate of suitable conditions and architecture technique has usually been taken into consideration to design appropriate electrochemical biosensors.Screen-printed carbon electrodes(SPCEs)have advantages such as simple and low cost fabrication as well as easy mass production.SPCEs simplify the use of electrochemical biosensors,avoid the fouling of electrodes due to their disposable character,and highlight the possibility of carrying out decentralized assays.SPCEs are more and more interesting because they can provide disposable-type chips for field applications.In recent years,SPCEs have been widely used to develop new electrochemical sensing platforms and improve the performance of the sensors.In this paper,we fabricated several disposable electrochemical biosensors based on SPCE substrate for determination of several biomacromolecules such as DNA,RNA and enzyme.The detailed materials are shown as follows:Chapter 1 generally introduced the principle and classification of electrochemical biosensor,SPCE and its surface modification,several immobilization techniques of bioactive element and three common electrochemical detection techniques.Then the recent advances in the applications of electrochemical biosensor based on SPCEs were reviewed.At last,the background of the dissertation and research contents were put forward.In chapter 2,an electrochemical DNA sensor was fabricated based on covalent immobilization of amino-terminated probe DNA onto the PDA modified SPCE.Immobilization was achieved via one-step Schiff base reaction between the amino group of the probe DNA and quinones in PDA.PDA was formed via electrochemical polymerization of dopamine monomer on the SPCE surface.After a sandwich-type hybridization reaction,the gold nanoparticle-labeled reporter DNA was bound onto the DNA sensor surface to further induce silver deposition in the presence of silver enhancer solutions.The electrochemical stripping signal of the deposited silver nanoparticles in KCl solution was used to monitor the hybridization reaction.Signal amplification enhanced the sensitivity for target DNA detection.The proposed method could detect target DNA at a linear range from 1.0 pM to 70 pM,with a detection limit of 0.3 pM.The DNA sensor exhibited good stability and acceptable reproducibility.The sensor also showed selectivity against non-complementary target DNA.In chapter 3,a sensitive and disposable electrochemical impedance RNA biosensor was fabricated based on DNA tetrahedron nanostructure probe and enzyme-catalyzed signal amplification reaction.The SPCE was modified with AuNPs through electrodeposition and subsequently functionalized with-SH of the DNA nanostructure.In the presence of the target microRNA,one part of the stem-loop structure of the DNA tetrahedron probe hybridized the target by opening the stem-loop structure and then another part of the probe hybridized to the reporter DNA with biotin label.At last,the avidin-HRP was attached to produce enzyme catalytic signal.HRP can catalyze CN oxidation to produce an insoluble precipitate accumulated on the electrode surface,which introduced a barrier to electron transfer between the redox probe in the electrolyte solution and the electrode,then caused the electron-transfer resistance of the sensor increased greatly.According to the above,signal amplification detection of the target microRNA can be carried out.In chapter 4,an electrochemical assay for the detection of PLA2 in serum was developed.The assay is composed of two major parts including MB-loaded liposomal nanoprobes and a bare SPCE.The probes were made by encapsulating the electroactive marker MB within nanometer-sized phospholipid liposomes.Due to low membrane permeability,the encapsulated MB exhibited no electrochemical signal.When the PLA2-containing sample and probe solution were mixed on the electrode surface,the phospholipids within the liposomal membrane were hydrolyzed by PLA2.The encapsulated MB was released,leading to the MB adsorption on the electrode surface.After washing,the SPCE with adsorbed MB was move into the blank buffer and the adsorbed MB was characterized by SWV.It was found that the peak current of SWV was linearly dependent on the PLA2 activity in the range of 5-200 U/L,and a detection limit as low as 3 U/L was obtained.This method provides the accurate assays of PLA2 in serum samples as well as the screening of PLA2 inhibitors,indicating its promising biomedical applications.In chapter 5,a simple method for the detection of PLA2 activity based on glucose-encapsulated liposomal nanoprobes and personal glucose meter(PGM)was presented.The probes were made by encapsulating the signal marker glucose within nanometer-sized phospholipid liposomes.Measurements were performed based on releasing encapsulated glucose from liposomes and subsequently detecting the released glucose using a commercial PGM.It was found that PGM signal was linearly dependent on the PLA2 activity.This assay opens another new avenue to bypassing the costly medical device development process.In the near future,it is expected that these sensors,in combination with aportable PGM,can be applied in POC testing.