A　new　sandwich-type　electrochemical　immunosensor　with　enhanced　sensitivity　was　developed　for　detection　of　alpha-fetoprotein　(AFP,　as　a　model　analyte)　in　biological　fluids　by　using　nanogold-enclosed　titania　nanoparticle　(AuTi)-labeled　secondary　antibody　on　a　gold-silver-graphene　hybrid　nanosheet　(AuAgGP)-functionalized　glassy　carbon　electrode　(GCE).　The　presence　of　the　AuAgGP　nanosheets　not　only　enhanced　the　immobilized　amount　of　biomolecules,　but　also　improved　the　electrochemical　properties　of　the　immunosensor.　With　the　aid　of　AuTi　nanolabels,　the　electrochemical　signal　was　greatly　amplified　in　comparison　with　pure　nanogold　or　titania-based　labels.　Under　optimal　conditions,　the　sensitivity　and　dynamic　range　of　the　immunosensor　were　evaluated　by　using　the　labeled　horseradish　peroxidase　on　the　AuTi　as　trace　and　H2O2　as　enzyme　substrate,　and　exhibited　a　wide　dynamic　range　of　0.001-200　ng　mL(-1)　with　a　low　detection　limit　(LOD)　of　0.5　pg　mL(-1)　AFP　(at　3　sigma).　Both　the　intra-　and　inter-assay　coefficients　of　variation　were　less　than　10%.　The　current　of　the　immunosensor　at　13th　day　was　as　much　as　90%　of　the　initial　current.　In　addition,　the　methodology　was　evaluated　for　8　positive　serum　specimens　obtained　from　hepatocarcinoma　patients　and　19　negative　sera,　and　validated　with　the　commercially　available　Roche　2010　Electrochemiluminescent　(ECL)　Automatic　Analyzer.　No　significant　differences　at　the　95%　confidence　level　were　encountered　between　two　methods.　(C)　2011　Elsevier　B.V.　All　rights　reserved.