Electrochemical Immunoassay For The Cancer Marker LMP-1 Using A Glassy Carbon Electrode Modified With Pd@Pt Nanoparticles And A Nanocomposite Consisting Of Graphene Sheets And Mwcnts

Objective:EBV(Epstein-Barr virus)is a human herpesvirus 4.The EBV is often a recessive infection in childhood and is prone to symptoms until the adult immune system is low.EBV relate to fatal infectious mononucleosis,Burkitt lymphoma,Hodgkin's disease,nasopharyngeal carcinoma,anaplastic large cell lymphoma,malignant histiocytosis and leiomyosarcoma.And a EBV genome has been detected in the majority of patients with nasopharyngeal carcinoma in southern China.EBV-derived latent membrane protein 1(LMP-1)is the only latent protein related cell transformation cancer gene confirmed currently,it can promote disease occurrence,development and metastasis by interfering with intracellular signal transduction pathway.Therefore,the sensitive detection of LMP-1 is essential for the early screening of related diseases.This study has constructed a new method of electrochemical immunosensor for detecting LMP-1 accurately,sensitively and rapidly.Methods:1.Synthesis of Pd@Pt bimetallic nanoparticles.All reactions were proceed under ultrasonic continuously for 4 h in water bath conditions,K2 Pt Cl4 and Na2 Pd Cl4 were reduced into Pd@Pt nanoparticles by AA,with the help of pluronic F127,Pd@Pt nanoparticles were dispersed uniformly.The bimetallic nanoparticles were characterized by TEM and XPS.2.With the aid of PDDA,Pd@Pt nanoparticles connected to the carboxylated MWCNTs by stiring overnight.Polyclonal antibody modified with Thi and HRP binding to MWCNTs via amide bond.Finally,the signal probe was prepared(Ab2/Thi/HRP/Pd@Pt-MWCNTs).It was characterized by SEM and electrochemical methods.3.GCE was used as the reaction substrate,and the PBS(0.1 M)with nitrogen gas was used as based reaction liquid.The p H value of the reaction liquid,the rate between graphene and MWCNTs,the antigen antibody incubation time and the concentration of hydrogen peroxide were optimized.Different concentrations of LMP-1 were detected by differential pulse voltammetry,and then standard curve was draw.Meanwhile,we evaluated the stability,sensitivity and selectivity of this immunosensor.Results:1.Pd@Pt bimetallic nanoparticles were characterized by TEM and XPS.The Pd@Pt nanoparticles were uniformly dispersed in the solution without obvious aggregation,and its' average size is 41.4 nm.Meanwhile,the XPS technique was used to further confirm the composition of the Pd@Pt nanoparticles.2.Under the optimum conditions,we detected the target LMP-1 with this prepared immunosensor.In the range from 1.00×10-2 to 4.00×101 ng m L-1 signal and the target concentration showed a linear correlation,R2=0.992,the limit of detection was 0.62 pg m L-1(S/N=3),the linear equation was I(u A)=7.803Log[C]+32.95.3.The sensor possess good analytical performance,selectivity and stability,and its' batch inter and intra difference were less than 5%.Conclusion:The surface of GCE was modified with the GS-MWCNTs composite,thereby increasing the electrode surface conductivity and the load of Ab1.This immunesensor could be used for rapid and sensitive detection of LMP-1.Based on immune response,and combined with the Pd@Pt bimetallic nanoparticles which have catalytic activity similar to HRP to catalyze H2O2,this immunosensor has a wide detection range,high specificity,good repeatability and high sensitivity,provides an effective method for clinical detection of LMP-1.