Electrochemical biosensor constructed using dual-signal amplification techniques (PER-HCR) for the ultrasensitive trace detection of ctDNA BRAF mutation.

The detection of BRAF mutations in circulating tumor DNA is beneficial for early diagnosis and personalized therapeutic approaches of melanoma. However, the sensitive and reliable trace detection of BRAF mutations remains a major challenge. Therefore, in this study, the combination of two isothermal signal amplification techniques (primer exchange reaction (PER) and hybridization chain reaction (HCR)) was proposed to construct an electrochemical biosensor for the efficient and sensitive detection of BRAF mutations. The presence of target triggered PER and HCR sequentially to achieve dual-cascade amplification of positive electrochemical signal feedback. Compared to the detection limit (1.27 pM or 0.23 pM) of single-signal amplification technology (PER or HCR), the detection limit of dual-signal amplification techniques was significantly lower (0.34 fM). In addition, the dual-signal amplification strategy exhibited other excellent sensing properties, such as a wide linear detection range (0.01 pM to 1 nM), excellent specificity, satisfactory reproducibility and stability, and the ability to achieve low-level target detection in diluted serum samples. Based on all of these features, the proposed strategy indicate broad application prospects for detecting low-abundance nucleic acid targets in clinical samples.