Engineering a G-quadruplex-based logic gate platform for sensitive assay of dual biomarkers of ovarian cancer

Abstract

As most single tumor markers have low sensitivity or specificity, the joint detection of multiple tumor biomarkers is helpful to improve the positive rate and specificity of early diagnosis. To establish a sensitive, specific, and rapid screening method for ovarian cancer, two enzyme-free logic gates were developed to realize the joint detection of carbohydrate antigen 125 (CA125) and carcinoembryonic antigen (CEA). G-quadruplex with peroxidase activity was used as a chromogenic catalyst in this colorimetric diagnosis. For NOR gate, in the absence of CEA and CA125, template DNA (Tem-DNA), CEA aptamer with half of the G-quadruplex sequence (CEA-apt), and CA125 aptamer with the other half of the G-quadruplex sequence (CA125-apt) form a double-stranded DNA equipped with one G-quadruplex structure which can strongly combine with hemin to obtain the peroxidase-like activity; here, the output of this NOR gate is set as 1, which could exclude the risk of ovarian cancer. For AND gate, CEA-apt and CA125-apt are separated from magnetic beads in the co-existence of the dual biomarkers. After adding Tem-DNA, the G-quadruplex structure constructs with hemin to exhibit the output as 1, presenting a high risk of ovarian cancer. Under the optimized condition, this novel assay can not only show a sharp color difference between physiological state and pathological state, but also be employed for the quantitative analysis of every single biomarker. The linear detection range and detection limit for CEA are 5–500 ng mL−1 and 0.88 ng mL−1, while they are for CA125 are 35–500 U mL−1 and 0.91 U mL−1, respectively. Moreover, the established approach was successfully applied in detecting CA125 and CEA in human serum samples, which provides great promise in the fields of clinical examination and disease diagnosis.