Sensitive immunosensor for cancer biomarker based on dual signal amplification strategy of graphene sheets and multienzyme functionalized carbon nanospheres

A novel electrochemical immunosensor for sensitive detection of cancer biomarker α-fetoprotein (AFP) is described that uses a graphene sheet sensor platform and functionalized carbon nanospheres (CNSs) labeled with horseradish peroxidase-secondary antibodies (HRP-Ab2). Greatly enhanced sensitivity for the cancer biomarker is based on a dual signal amplification strategy: first, the synthesized CNSs yielded a homogeneous and narrow size distribution, which allowed several binding events of HRP-Ab2 on each nanosphere. Enhanced sensitivity was achieved by introducing the multibioconjugates of HRP-Ab2-CNSs onto the electrode surface through "sandwich" immunoreactions. Second, functionalized graphene sheets used for the biosensor platform increased the surface area to capture a large amount of primary antibodies (Ab1), thus amplifying the detection response. On the basis of the dual signal amplification strategy of graphene sheets and the multienzyme labeling, the developed immunosen-sor showed a 7-fold increase in detection signal compared to the immunosensor without graphene modification and CNSs labeling. The proposed method could respond to 0.02 ng mL^-1 AFP with a linear calibration range from 0.05 to 6 ng mL^-1. This amplification strategy is a promising platform for clinical screening of cancer biomarkers and point-of-care diagnostics.