循环肿瘤 DNA (ctDNA) 是早期肿瘤检测的关键生物标志物。然而，准确定量人血清中的低丰度 ctDNA 仍然是一个重大挑战。为了应对这一挑战，我们推出了一种双峰生物传感器，专门用于检测特定非小细胞肺癌 (NSCLC) 患者的表皮生长因子受体 (EGFR) 突变 L858R。该生物传感器利用双 CRISPR-Cas12a 系统通过荧光和电化学信号量化目标。在我们的系统中，EGFR L858R 对限制酶 MscI 的消化表现出抗性，MscI 会激活第一个 CRISPR-Cas12a 蛋白，并抑制磁珠与荧光素 (FAM) 标记的杂交链式反应 (HCR) 产物的结合，从而减少荧光信号。这种激活还抑制第二个 CRISPR-Cas12a 蛋白的裂解活性，使电极能够维持来自纳米材料的更高的电化学信号。野生型 EGFR (wt EGFR) 产生相反的效果。因此，可以使用荧光和电化学信号准确定量和验证 EGFR L858R 的浓度。该生物传感器提供 10 fM 至 1 μM 的动态检测范围，检测限为 372 aM。它表现出出色的特异性、重现性、稳定性和回收率。此外，该传感器增强的分析灵敏度凸显了其在生物传感应用和早期疾病诊断中的关键作用。

Circulating tumor DNA (ctDNA) is a critical biomarker for early tumor detection. However, accurately quantifying low-abundance ctDNA in human serum remains a significant challenge. To address this challenge, we introduce a bimodal biosensor tailored for detecting the epidermal growth factor receptor (EGFR) mutation L858R in specific nonsmall cell lung cancer (NSCLC) patients. This biosensor utilizes dual CRISPR-Cas12a systems to quantify the target via fluorescence and electrochemical signals. In our system, the EGFR L858R exhibits resistance to digestion by the restriction enzyme MscI, which activates the first CRISPR-Cas12a protein and inhibits the binding of magnetic beads with fluorescein (FAM)-labeled hybridization chain reaction (HCR) products, thereby reducing the fluorescence signal. This activation also inhibits the cleavage activity of the second CRISPR-Cas12a protein, allowing the electrode to sustain a higher electrochemical signal from nanomaterials. The wild-type EGFR (wt EGFR) produces the opposite effect. Consequently, the concentration of EGFR L858R can be accurately quantified and verified using both fluorescence and electrochemical signals. The biosensor offers a dynamic detection ranging from 10 fM to 1 μM, with a detection limit of 372 aM. It demonstrates excellent specificity, reproducibility, stability, and recovery rates. Moreover, the sensor's enhanced analytical sensitivity highlights its critical role in biosensing applications and early disease diagnosis.