8-氧鸟嘌呤（8-oxoG）基DNA损伤是DNA损伤中最常见的类型，对基因表达有很大影响。因此，准确量化8-oxoG基DNA损伤在临床意义上具有很高的重要性。然而，目前用于8-oxoG检测的方法很难在方便性、低成本和灵敏度之间取得平衡。在此，我们提出并研究了CRISPR-Cas12a系统中缩短的crRNA模式，并大大增强了其信噪比。利用缩短的crRNA模式的优势，我们进一步开发了一种CRISPR增强结构切换适配体测定法（CESA）用于检测8-oxoG。通过检测游离8-oxoG和基因组DNA上的8-oxoG，对CESA的分析性能进行了深入研究。CESA对游离8-oxoG显示出了令人印象深刻的敏感性，检测限和定量限分别为32.3 pm 和0.107 nM。当检测基因组DNA上的8-oxoG时，这些限制略有增加，分别为64.5 pm和0.215 nM。为了证明CESA系统在临床可行性和重要性方面的作用，我们进一步将其应用于衡量7份血浆样本中的8-oxoG水平（宫颈癌，11.87 ± 0.69 nM 对照组，2.66 ± 0.42 nM）、24份血浆样本中的8-oxoG水平（无精子症，22.29 ± 7.48 nM 正常精子，9.75 ± 3.59 nM）、10份乳腺组织基因组DNA样本中的8-oxoG水平（乳腺癌，2.77 ± 0.63 nM/μg 对照组，0.41 ± 0.09 nM/μg）以及24份精子基因组DNA样本中的8-oxoG水平（无精子症，28.62 ± 4.84 正常精子，16.67 ± 3.31）。此项工作不仅提出了一种基于缩短crRNA设计范式的CRISPR-Cas12a基生物传感器开发方法，还为检测8-oxoG基DNA损伤提供了强大的工具。Copyright © 2023 Elsevier B.V. All rights reserved.

8-oxoguanine (8-oxoG) based DNA damage is the most common type of DNA damage which greatly affect gene expression. Therefore, accurate quantification of 8-oxoG based DNA damage is of high clinical significance. However, current methods for 8-oxoG detection struggle to balance convenience, low cost, and sensitivity. Herein, we have proposed and investigated the shortened crRNA mode of CRISPR-Cas12a system and greatly enhanced its signal-to-noise ratio. Taking advantages of the shortened crRNA mode, we further developed a CRISPR-enhanced structure-switching aptamer assay (CESA) for 8-oxoG. The analytical performance of CESA was thoroughly investigated via detecting free 8-oxoG and 8-oxoG on gDNA. The CESA displayed impressive sensitivity for free 8-oxoG, with detection and quantification limits of 32.3 pM and 0.107 nM. These limits modestly rose to 64.5 pM and 0.215 nM when examining 8-oxoG on gDNA. To demonstrate the clinical practicability and significance of the CESA system, we further applied it to measuring 8-oxoG levels in 7 plasma samples (Cervical carcinoma, 11.87 ± 0.69 nM VS. Healthy control, 2.66 ± 0.42 nM), 24 seminal plasma samples (Asthenospermia, 22.29 ± 7.48 nM VS. Normal sperm, 9.75 ± 3.59 nM), 10 breast-tissue gDNA samples (Breast cancer, 2.77 ± 0.63 nM/μg VS. Healthy control, 0.41 ± 0.09 nM/μg), and 24 sperm gDNA samples (Asthenospermia, 28.62 ± 4.84 VS. Normal sperm, 16.67 ± 3.31). This work not only proposes a novel design paradigm of shortened crRNA for developing CRISPR-Cas12a based biosensors but also offers a powerful tool for detecting 8-oxoG based DNA damage.Copyright © 2023 Elsevier B.V. All rights reserved.