单位:[1]Department of Gynecological Oncology,National Clinical Research Center for Obstetrics and Gynecology,Cancer Biology Research Center (Key Laboratory of the Ministry of Education),Tongji Hospital,Tongji Medical College,Huazhong University of Science and Technology,Wuhan 430070,China.妇产科学系科研平台肿瘤生物医学中心华中科技大学同济医学院附属同济医院肿瘤科[2]Department of Obstetrics and Gynecology, The First Affiliated Hospital of USTC, Division of Life Sciences and Medicine, University of Science and Technology of China, Hefei 230001, China.[3]State Key Laboratory of Magnetic Resonance and Atomic Molecular Physics, Key Laboratory of Magnetic Resonance in Biological Systems, National Center for Magnetic Resonance in Wuhan, Wuhan Institute of Physics and Mathematics, Innovation Academy for Precision Measurement Science and Technology, Chinese Academy of Sciences - Wuhan National Laboratory for Optoelectronics, Wuhan 430071, China.[4]Department of Laboratory Medicine, Maternal and Child Health Hospital of Hubei Province, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430070, China.[5]Optics Valley Laboratory, Hubei 430074, China.
Timely identification of human papillomavirus (HPV) infection is crucial for the prevention of cervical cancer. Current HPV detection methods mainly rely on polymerase chain reaction (PCR), which often requires bulky equipment and a long assay time. In this work, we report a heating-membrane-assisted multiplexed microfluidics platform that couples recombinase polymerase amplification (RPA) and CRISPR technology (termed M3-CRISPR) for fast and low-cost detection of multiple HPV subtypes. The heating membrane can provide convenient temperature control for the on-chip RPA and CRISPR assays. This stand-alone system allows simultaneous detection of HPV16 and HPV18 with high specificity and detection sensitivity (0.5 nM and 1 × 10-18 M for unamplified and amplified plasmids, respectively) in 30 min with a fluorescence-based readout. Furthermore, we introduced an optimized lateral flow dipstick (LFD) into the portable system to allow visualized detection of HPV DNA. The LFD-based readout also reached a detection sensitivity of 1 × 10-18 M for amplified plasmids and realized successful detection of HPV subtypes in the clinical samples. Finally, we established an automatic microfluidic system that enables the sample-in-answer-out detection of HPV subtypes. We believe that this fast, convenient, and affordable molecular diagnostic platform can serve as a useful tool in point-of-care testing of HPV or other pathogens.
基金:
National Natural Science Foundation of China (22174150,
22074152), the Chinese Academy of Sciences (Y9Y1041001,
YJKYYQ20170026), and the Fundamental Research Funds for
the Central Universities (WK9110000176).
第一作者单位:[1]Department of Gynecological Oncology,National Clinical Research Center for Obstetrics and Gynecology,Cancer Biology Research Center (Key Laboratory of the Ministry of Education),Tongji Hospital,Tongji Medical College,Huazhong University of Science and Technology,Wuhan 430070,China.[2]Department of Obstetrics and Gynecology, The First Affiliated Hospital of USTC, Division of Life Sciences and Medicine, University of Science and Technology of China, Hefei 230001, China.
通讯作者:
通讯机构:[1]Department of Gynecological Oncology,National Clinical Research Center for Obstetrics and Gynecology,Cancer Biology Research Center (Key Laboratory of the Ministry of Education),Tongji Hospital,Tongji Medical College,Huazhong University of Science and Technology,Wuhan 430070,China.[3]State Key Laboratory of Magnetic Resonance and Atomic Molecular Physics, Key Laboratory of Magnetic Resonance in Biological Systems, National Center for Magnetic Resonance in Wuhan, Wuhan Institute of Physics and Mathematics, Innovation Academy for Precision Measurement Science and Technology, Chinese Academy of Sciences - Wuhan National Laboratory for Optoelectronics, Wuhan 430071, China.[*1]State Key Laboratory of Magnetic Resonance and Atomic Molecular Physics, Key Laboratory of Magnetic Resonance in Biological Systems, National Center for Magnetic Resonance in Wuhan, Wuhan Institute of Physics and Mathematics, Innovation Academy for Precision Measurement Science and Technology, Chinese Academy of Sciences − Wuhan National Laboratory for Optoelectronics, Wuhan 430071, China[*2]Department of Gynecological Oncology,National Clinical Research Center for Obstetrics and Gynecology,Cancer Biology Research Center (Key Laboratory of the Ministry of Education),Tongji Hospital,Tongji Medical College,Huazhong University of Science and Technology,Wuhan 430070,China
推荐引用方式(GB/T 7714):
Zhou Hu,Xu Zhichen,He Liang,et al.Coupling CRISPR/Cas12a and Recombinase Polymerase Amplification on a Stand-Alone Microfluidics Platform for Fast and Parallel Nucleic Acid Detection[J].ANALYTICAL CHEMISTRY.2023,95(6):3379-3389.doi:10.1021/acs.analchem.2c04713.
APA:
Zhou Hu,Xu Zhichen,He Liang,Wang Zhijie,Zhang Tao...&Huang Xiaoyuan.(2023).Coupling CRISPR/Cas12a and Recombinase Polymerase Amplification on a Stand-Alone Microfluidics Platform for Fast and Parallel Nucleic Acid Detection.ANALYTICAL CHEMISTRY,95,(6)
MLA:
Zhou Hu,et al."Coupling CRISPR/Cas12a and Recombinase Polymerase Amplification on a Stand-Alone Microfluidics Platform for Fast and Parallel Nucleic Acid Detection".ANALYTICAL CHEMISTRY 95..6(2023):3379-3389