Preparation Of Polymer/Graphene Oxide Composites For The High Sensitivity Detection Of Tumor Markers

Cancer is one of the major diseases that threaten human life in the current society.The detection of tumor marker?TM?is the only effective method for early detection of asymptomatic microscopic tumors.Electrochemical immunosensor?EI?is a novel biosensor that combines immunoassay and electrochemical sensing.It has both the high specificity of immunoassay and the high sensitivity of electronic analytical chemistry.However,the extremely low TM content of early oncology patients presents great difficulties for detection and screening.Therefore,by improving the detection sensitivity of tumor marker signal amplification,building high performance EI is the key to our research.In this paper,we have constructed a variety of electrochemical immunosensing platforms that enable high sensitivity and high specificity detection of one or more TMs.By synthesizing a variety of polymer/graphene oxide nanocomposites,these sandwich immunosensor successfully achieved the exponential amplification of the signal value by detecting TMs.The content of the thesis is mainly divided into three parts.1.An electrochemical immunosensor for highly sensitive detection of the tumor marker AFP has been successfully developed.Specifically,poly?2-hydroxyethyl methacrylate?-r-poly?acrylic acid??PHEMA-r-PAA?with a large number of Active carboxyl group and hydroxyl group was synthesized by RAFT polymerization using HEMA and AA as monomers.The active hydroxyl group on the main chain provides a large number of sites for the loading of the signal molecule Aq,and the carboxyl group was used to bond GO to construct the nanocomposite.The nanocomposites constructed by PHEMA-r-PAA and GO were used for the preparation of electrochemical immunosensors and achieve highly sensitive detection of AFP.The wide range of detection was achieved at 3.5 pg/mL-35 ng/mL and the extremely low detection limit of 106 fg/mL.2.An electrochemical immunosensor capable of simultaneously detecting three tumor markers?PSA,AFP,CA 125?has been developed.Poly?styrene-alt-maleic anhydride??PSMA?was prepared by a thermally initiated free radical polymerization using St and MAH as monomers while controlling the molar ratio therebetween.After hydrolysis with ammonia water,three signal molecules Aq,Fc,ASA were respectively loaded on it.Subsequently,the amino-modified GO was loaded with three different detection antibodies Ab₂?anti-PSA,anti-AFP,anti-CA 125?after activation of the carboxyl group.Finally,the polymer loaded with the three signal molecules was grafted to obtain the nanocomposites probe with signal amplification capability?PSMA-Aq/GO-Ab₂,PSMA-Fc/GO-Ab₂,PSMA-ASA/GO-Ab₂?.The simultaneous detection of tumor markers is achieved by specific binding between antigen-antibodies.Under the optimized conditions,the multiplexed immunosensor displayed an excellent linear response in the range of 1.13 pg/mL-113 ng/mL for PSA,0.35 pg/mL-35ng/mL for AFP and 0.025 U/mL-250 U/mL for CA 125.The detection limit was 86fg/mL,14 fg/mL and 0.0019 U/mL for PSA,AFP and CA 125,respectively,which well illustrates the high sensitivity of our platform.3.A novel electrochemical immunoassay platform for multi-signal amplification has been constructed and applied to the simultaneous detection of three tumor markers.The platform was based on the development of polyethyleneimine?PEI?-polydopamine?PDA?/reduced graphene oxide?rGO?nanocomposites.Among which,PEI was used to immobilize different signal labels,and PDA/rGO,the substrates of nanoprobes,can be loaded with large amounts of PEI and detection antibodies?Ab₂?simultaneously.The tumor markers of carcinoembryonic antigen?CEA?,prostate specific antigen?PSA?and human a-fetoprotein?AFP?were employed as model test molecules.Ferrocenecarboxylic acid?Fc?,anthraquinone-2-carboxylic acid?Aq?and acetylsalicylic acid?ASA?were employed as distinguishable signal tags.Electrochemical responses of as-prepared sandwich-type immunoassay were investigated by cyclic voltammetry?CV?and differential pulse voltammetry?DPV?.Under the optimized conditions,the multiplexed immunoassay displayed excellent linear response in the range of 0.102 pg/mL-112 ng/mL for CEA and PSA,0.117pg/mL-117 ng/mL for AFP.The detection limit was 91 fg/mL,101 fg/mL and 83fg/mL for CEA,PSA and AFP,respectively.Moreover,this assay provides good reproducibility and stability,ensuring a promising future in simultaneous immunoassay of multiple protein biomarkers.