We report a sandwich-type electrochemical immunoassay for simultaneous determination of carcinoembryonic antigen (CEA) and alpha-fetoprotein (AFP), using multifunctionalized gold nanoparticles as signal tags. Two nanotags including thionine-Au nanoparticles and ferrocene-Au nanoparticles were synthesized and used as distinguishable indicators. The sensor was fabricated by means of co-immobilization of primary anti-CEA (AbCEA) and anti-AFP (AbAFP) antibodies on the surface of Fe3O4 nanoparticle, the strategy is based on observation distinct voltammetric peak derived from distinguish probe (AbCEA-Au-thionine and AbAFP-Au -ferrocene nanotages). Experimental results revealed that this immunosensor enabled simultaneous determination of CEA and AFP in the linear ranges of (0.05- 120 ngmL-1) and (0.05-100 ngmL-1), respectively. The detection limits (DL) for AFP and CEA were 0.012 ngmL-1 and 0.018 ngmL-1, respectively (S/N=3). Furthermore, the methodology was also evaluated for the analysis of AFP and CEA in the clinical human serum specimens, the obtained results was in desirable agreement with clinical data.
