DNA detection plays an important role in the rapid screening of cancers and early diagnosis of infectious diseases. Here, we developed a simple, versatile, electric field-enhanced (EFE), electrochemical CRISPR biosensor to detect DNA targets in a homogeneous solution phase. To improve the detection sensitivity, we applied a pulsed electric field to enrich nucleic acids on the electrode surface. The EFE electrochemical CRISPR biosensor takes advantage of the diffusivity difference between electrochemical oligonucleotide probes and CRISPR-cleaved probes toward a negatively charged working electrode, enabling simple and sensitive electrochemical detection of DNA without the need for complicated immobilization processing of electrochemical probes. Our developed CRISPR biosensor directly detects unamplified human papillomavirus-16 (HPV-16) DNA with a sensitivity of 1 pM. Further, the EFE electrochemical CRISPR biosensor coupled with recombinase polymerase amplification successfully detects HPV-16 DNA in clinical samples. Thus, the EFE electrochemical CRISPR biosensor provides a simple, robust, and sensitive detection method for nucleic acid-based molecular diagnostics.

DNA检测在癌症的快速筛查和传染病的早期诊断中发挥着重要作用。在这里，我们开发了一种简单、多功能、电场增强 (EFE) 的电化学 CRISPR 生物传感器，用于检测均质溶液相中的 DNA 靶标。为了提高检测灵敏度，我们应用脉冲电场富集电极表面的核酸。EFE 电化学 CRISPR 生物传感器利用电化学寡核苷酸探针和 CRISPR 切割探针之间朝向带负电的工作电极的扩散性差异，实现简单、灵敏的 DNA 电化学检测，无需对电化学探针进行复杂的固定化处理。我们开发的 CRISPR 生物传感器可直接检测未扩增的人乳头瘤病毒 16 (HPV-16) DNA，灵敏度为 1 pM。此外，EFE 电化学 CRISPR 生物传感器与重组酶聚合酶扩增相结合，成功检测临床样本中的 HPV-16 DNA。因此，EFE电化学CRISPR生物传感器为基于核酸的分子诊断提供了一种简单、稳健且灵敏的检测方法。