Clustered Regularly Interspaced Short Palindromic Repeats-Mediated Amplification-Free Detection of Viral DNAs Using Surface-Enhanced Raman Spectroscopy-Active Nanoarray
Clustered Regularly Interspaced Short Palindromic Repeats-Mediated Amplification-Free Detection of Viral DNAs Using Surface-Enhanced Raman Spectroscopy-Active Nanoarray
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DOI：
10.1021/acsnano.1c03975
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发表时间：
2021-08-09
期刊：
影响因子：
17.1
通讯作者：
Choi, Jeong-Woo
中科院分区：
文献类型：
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作者：
Choi, Jin-Ha;Shin, Minkyu;Choi, Jeong-Woo
核酸生物标志物已被广泛用于检测各种病毒相关疾病，包括最近的大流行性COVID-19。基于CRISPR-Cas的反式激活现象已经显示出开发核酸的灵敏和选择性检测的优异潜力。然而，当需要灵敏和选择性地监测靶核酸时，通常需要核酸扩增步骤。为了克服上述挑战，我们通过表面增强拉曼光谱（Sers）辅助的超灵敏检测系统开发了一种基于CRISPR-Cas 12 a的无核酸扩增生物传感器。我们将通过病毒DNA激活的CRISPR-Cas 12 a与由ssDNA固定的拉曼探针功能化的Au纳米颗粒（RAuNPs）组成的拉曼敏感系统整合在氧化石墨烯（GO）/三角形Au纳米花阵列上。使用这种基于CRISPR的拉曼敏感系统提高了多病毒DNA的检测灵敏度，如B肝炎病毒（HBV），人乳头瘤病毒16（HPV-16）和HPV-18，具有极低的检测限和从1 aM到100 pM的广泛检测范围，无需扩增步骤。我们认为，这种超灵敏的无扩增核酸检测系统可以广泛应用于病毒感染，癌症和几种遗传疾病的精确和早期诊断。
Nucleic acid biomarkers have been widely used to detect various viral-associated diseases, including the recent pandemic COVID-19. The CRISPR-Cas-based trans-activating phenomenon has shown excellent potential for developing sensitive and selective detection of nucleic acids. However, the nucleic acid amplification steps are typically required when sensitive and selective monitoring of the target nucleic acid is needed. To overcome the aforementioned challenges, we developed a CRISPR-Cas12a-based nucleic acid amplification-free biosensor by a surface-enhanced Raman spectroscopy (SERS)-assisted ultrasensitive detection system. We integrated the activated CRISPR-Cas12a by viral DNA with a Raman-sensitive system composed of ssDNAimmobilized Raman probe-functionalized Au nanoparticles (RAuNPs) on the graphene oxide (GO)/triangle Au nanoflower array. Using this CRISPR-based Raman-sensitive system improved the detection sensitivity of the multiviral DNAs such as hepatitis B virus (HBV), human papillomavirus 16 (HPV-16), and HPV-18 with an extremely low detection limit and vast detection range from 1 aM to 100 pM without the amplification steps. We suggest that this ultrasensitive amplification-free detection system for nucleic acids can be widely applied to the precise and early diagnosis of viral infections, cancers, and several genetic diseases.