Graphene And Chitosan Nanocomplexes Were Used To Construct Dual-target Tumor Marker Current-type Immunosensors

Since most cancer biomarkers are not specific to a particular cancer, single-analyte detection might lead to false positives in cancer diagnosis. Thus, the simultaneous detection of multiplexed biomarkers gained great value. Amperometric electrochemical immunoassay has shown its unique advantages due to its low cost, fast analysis and simple instruments. Recently, an improved sensitivity in multiplexed detection has been obtained by coupling of electrochemical immunoassay with nanomaterials. With the sandwich-format immunoassay, we constructed the electrochemical immunosensors for simultaneous detection of carcinoembryonic antigen (CEA) and alpha-fetoprotein (AFP), based on carboxyl graphene nanosheets (CGS), chitosan (CHIT), Au nanoparticles (AuNPs), toluidine blue (TB), Prussian blue (PB), ferrocenecarboxylic acid (Fc). This work got the following achievements:1. Electrochemical immunosensor for simultaneous detection of CEA and AFP based on CGS-TB and CGS-PB nanocomposites. CGS can absorb enormous TB and PB molecules due to its large surface aera, following by immobilizing antibodies, the nanoprobes were obtained. The excellent membrane-formability of CHIT and the good conductivity of AuNPs make CHIT-AuNPs a good substrate for immobilize anti-CEA and anti-AFP. With the cooperation of CGS, CHIT and AuNPs, the immunosensor displayed a good result. A linear range of0.5ng mL-1to60ng mL-1for both analytes with the detection limit of0.1ng mL-1for CEA and0.05ng mL-1for AFP was obtained. The assay results of serum samples with the proposed method were in a good agreement with the reference values from standard ELISA method.2. Electrochemical immunosensor for simultaneous detection of CEA and AFP based on CHIT-PB-AuNPs and CHIT-Fc-AuNPs nanocomposites. CHIT is an excellent carrier with large amounts of amino groups and good biocompatibility. After immobilized TB and PB, it can still absorb substantial AuNPs, which can further absorbed antinbodies to form signal tags. As a good immunosensing platform, CHIT-AuNPs modified electrodes were used to immobilize capture anti-AFP and anti-CEA. The immunosensor revealed a good performance with a wide range of0.05-100ng mL-1for both AFP and CEA. The detection limits was0.03ng mL-1for AFP and0.02ng mL-1for CEA. The assay results of serum samples with the proposed method were in a good agreement with the reference values from standard ELISA method.