Fabrication of electrochemical biosensor composed of multi-functional DNA 4 way junction for TNF-α detection in human serum

Tumor necrosis factor (TNF-α) is a representative cytokine family known to induce multiple signaling cascades leading to various cellular responses, such as cell death, survival, and differentiation. It has been reported that blocking the action of TNF-α in various diseases can improve disease prognosis. Therefore, it is important to monitor TNF-α in patient plasma and properly regulate its action. In this study, we report a label-free electrochemical biosensor consisting of a multifunctional DNA 4-way junction (MF-4WJ) for TNF-α detection in human serum. MF-4WJ does not require additional labeling and signal amplification processes. The electrochemical properties of functionalized MF-4WJ were evaluated by cyclic voltammetry (CV) in the presence of Ag⁺ intercalated between the mismatched sequences of MF-aptamers as redox-active species. Afterward, CV was carried out to evaluate the performance of the fabricated biosensor. The proposed label-free electrochemical biosensor was able to effectively detect TNF-α in a dynamic range of 0.15 pg/ml to 150 ng/ml. Limit of detection (LOD) was at 0.07 pg/ml in HEPES. Moreover, it was confirmed that even in 10% diluted human serum, TNF-α could be detected in an excellent dynamic range of 0.15 pg/ml to ∼ 15 ng/ml and LOD was at 0.14 pg/ml in 10% diluted human serum.