Anti‐CRISPR Protein Regulates CRISPR/Cas12a Fusogenic‐Nanovesicle‐Based Platform for Extracellular Vesicle‐Encapsulated Non‐Nucleic Acid Target In‐Vesicle Detection
Abstract
ABSTRACT The detection of non‐nucleic acid targets encapsulated in extracellular vesicles (EVs) faces two major challenges: (1) difficulties in efficient isolation and the risk of content degradation, and (2) the low abundance of target molecules encapsulated in EVs always leads to failed signal transduction and inadequate output signal intensity. To overcome these limitations, we propose a high‐efficiency in‐vesicle analysis strategy that integrates targeting probe delivery and regulation by protein signal amplification. By applying aptamer‐mediated membrane fusion and “locked‐activated” CRISPR‐Cas12a‐AcrVA1 (LACA) for protein signal regulation, we fabricated a yly12‐aptamer‐functionalized self‐assembled nanovesicle which encapsulate LACA‐system (yly12‐lipo@Cas12a nanovesicle) as an in‐vesicle bioanalytical platform. Leveraging the high specificity of the aptamer and the regulatory function of AcrVA1 in selectively modulating Cas12a activity, the platform enables highly specifiec and sensitive detection, offering advantages of simple operation and versatility across platforms within only 2.5 h. Clinical analysis demonstrated effective differentiation between patients and healthy controls, yielding high diagnostic performance with an AUC of 0.965. The proposed platform shows great potential for EV‐carrying protein biomarker analysis and has broad prospects for the disease's diagnosis in clinical settings.
Article Details
Authors (10)
Qiaoxuan Zhang
Department of Laboratory Medicine/State Key Laboratory of Dampness Syndrome of Chinese Medicine The Second Affiliated Hospital of Guangzhou University of Chinese Medicine Guangzhou China
Yaling Li
State Key Laboratory of Green Pesticide, College of Chemistry
Yuanzhe Li
Liqiao Han
Department of Laboratory Medicine/State Key Laboratory of Dampness Syndrome of Chinese Medicine The Second Affiliated Hospital of Guangzhou University of Chinese Medicine Guangzhou China
Jun Yan
School of Materials Science and Engineering
Min Zhan
Department of Laboratory Medicine/State Key Laboratory of Dampness Syndrome of Chinese Medicine The Second Affiliated Hospital of Guangzhou University of Chinese Medicine Guangzhou China
Ting Liu
Peifeng Ke
Department of Laboratory Medicine/State Key Laboratory of Dampness Syndrome of Chinese Medicine The Second Affiliated Hospital of Guangzhou University of Chinese Medicine Guangzhou China
Qiqin Wang
Institute of Pharmaceutical Analysis College of Pharmacy/State Key Laboratory of Bioactive Molecules and Draggability Assessment/International Cooperative Laboratory of Traditional Chinese Medicine Modernization and Innovative Drug Development of Ministry of Education (MOE) of China Jinan University Guangzhou China
Xianzhang Huang
Department of Laboratory Medicine/State Key Laboratory of Dampness Syndrome of Chinese Medicine The Second Affiliated Hospital of Guangzhou University of Chinese Medicine Guangzhou China