Detection of carcinoembryonic antigen and α-fetoprotein exploiting a 3D DNA walker strategy

An electrochemical immunosensor was developed for simultaneous detection of two biomarkers. The combination of DNAzyme, DNA walker machine and nicking enzyme strategy enhanced the sensitivity of the immunosensor. The utilization of Y-junction DNA structure promoted the catalytic efficiency of Nt.BbvCI, and strengthened the stability of the immunosensor. The intrinsic recognition and conjugation between DNA complementary strands and antigen/antibody ensured the high selectivity of the immunosensor. The highly sensitivity composed of pg mL−1 level of LOD and six orders of magnitude of linear range. Carcinoembryonic antigen (CEA) and α-fetoprotein (AFP) are known cancer biomarkers and are widely used for diagnosis in clinical research. Here, we report an Mg2+-based DNAzyme-mediated three-dimension (3D) DNA walker strategy and demonstrated its use for the detection of CEA and AFP. The 3D DNA walker constructed 3D tracks using hollow carbon nanospheres (HCS) as a support. In the presence of the biomarkers and the Mg2+ cofactor, the DNA walker actively generated multiple copies of intermediate DNA (iDNA) fragments from the substrate probe that is immobilized on the HCS support. With the use of a nicking enzyme, amplification is triggered by iDNA, resulting in the decrease of the redox signal of methylene blue (MB) and ferrocene (Fc). For the detection of CEA, the immunosensor has a limit of detection of 0.3 pg mL−1 with a linear range between 1 pg mL−1 and 1000 ng mL−1. For AFP detection, the limit of detection is 0.5 pg mL−1 with a linear range of 1 pg mL−1 to 100 ng mL−1. As part of this study, the employment of this strategy in serum samples is demonstrated for the detection of CEA and AFP.