Study Of Building Protein Kinases And Arsenite Electrochemical Luminescence Sensor

In this dissertation,two novel electrochemical luminescence（ECL）sensors have been constructed combined with nanomaterials,which are successfully used in the detection of protein kinase A and arsenite.The main contents are summarized as follows:1.The introduction part mainly summarizes the development,characteristics and application in biological detection and environmental testing of ECL.Then the nature of the gold nanoparticles（Au NPs）and graphite carbon nitride nanosheets and its application in ECL were introduced.And the significance of detecting protein kinase A and arsenite and the present main testing methods were introduced.Finally,the main research work and practical application of this paper are introduced.2.A novel ECL biosensor using gold nanoparticles as a signal amplification probe of S₂O₈^2--O₂ system was developed for ultrasensitive detecting of kinase activity and inhibition.Peptides were modified with chitosan on the surface of GCE.In the presence of adenosine 5′-[γ-thio]triphosphate（ATP）cosubstrate and protein kinase A（PKA）,peptides would be phosphorylated and Au NPs were bind onto the phosphorylated peptides coupled on the surface of electrode through Au-S bond.The ECL signal of S₂O₈^2--O₂ system could be enhanced by closely contacted Au NPs.The ECL enhancement was positively correlated with the PKA activity.Therefore,on the basis of the Au NPs enhanced effect,the activity of protein kinases was detected with ultrahigh sensitivity,low detection limit and simple process.This biosensor is also successfully used for quantitative analysis of PKA activity in serum samples and qualitative screening of kinase inhibition,manifesting that the method shows a good potential application in pharmaceutical research and clinical diagnosis.3.A ratiometric ECL indicator for ultratrace arsenite（As（Ⅲ））detection was constructed.We modified gold functional carbon nitride nanosheets（Au-g-C₃N₄ NSs）on the surface of GCE and the Ars-3 aptamer would connect with Au NPs of Au-g-C₃N₄ NSs by Au-S bond.The electropositive cationic polymer PDDA can be combined with a negatively charged adaptive phosphate backbone of Ars-3 aptamer by electrostatic interaction.As（Ⅲ）would be specifically combined with Ars-3aptamer which would lead to the formation change of Ars-3 aptamer.So that Ars-3aptamer would separate from PDDA and absorb more positive charge Ru（bpy）₃²⁺by electrostatic action.The As（Ⅲ）detection based on quenching ECL of Au-g-C₃N₄NSs by As（0）and Ru（bpy）₃²⁺through electron transfer（ET）and resonance energy transfer（RET）,respectively,and meanwhile generating an ECL of Ru（bpy）₃²⁺with increased intensity.Compared to ECL quenching of Au-g-C₃N₄ NSs simplex by As（0）（linearity range of 1-50 ppb,LOD of 0.62 ppb）,the sensitivity and selectivity for detecting As（Ⅲ）is vastly enhanced by dual quenching effects of As（0）coupled with Ru（bpy）₃²⁺and the corresponding second ECL signal of Ru（bpy）₃²⁺.The logarithmic value of ECL₆₂₀/ECL₄₆₀ is linearly proportional to the logarithm of As（Ⅲ）concentration from 0.001 ppt to 10 ppb.The LOD is estimated to be 0.0007 ppt.Also,this ratiometric ECL indicator has been successfully used for analysis of natural water samples with satisfactory results.