Rapid Discrimination of Single‐Virus Entry Pathways via a Bioorthogonal Activatable Fluorescent Probe
Abstract
ABSTRACT Viral transmembrane entry is a critical yet heterogeneous process that governs infectivity and pathogenesis. However, directly distinguishing virus attachment from internalization and resolving distinct entry mechanisms in real time at the single‐virus level remains challenging. Here, we introduce a Bioorthogonal Click‐Activatable Viral Labeling (CAVL) system for spatiotemporally resolved visualization of viral entry dynamics. This approach combines a membrane‐anchored, tetrazine‐quenched BODIPY probe (BDP‐Tz‐MAL) with viral particles dually labeled with trans‐cyclooctene (TCO) on their surface and quantum dots (QDs) within their core. Virus–cell contact triggers a rapid inverse electron‐demand Diels–Alder (iEDDA) reaction, generating a turn‐on fluorescence signal exclusively during transmembrane entry. By integrating this activatable signal with a persistent internal QDs label, we achieve real‐time discrimination of endocytic and non‐endocytic entry pathways at single‐particle resolution. The CAVL platform operates without genetic modification, applies to diverse enveloped and non‐enveloped viruses, and provides a quantitative, background‐suppressed strategy for elucidating viral entry mechanisms across broad biological contexts.
Article Details
Authors (10)
Ai‐Xin Ma
State Key Laboratory of Medicinal Chemical Biology Tianjin Key Laboratory of Biosensing and Molecular Recognition Frontiers Science Centre For New Organic Matter Research Centre for Analytical Sciences College of Chemistry School of Medicine and Frontiers Science Center for Cell Responses Nankai University Tianjin P.R. China
Zhi‐Gang Wang
State Key Laboratory of Medicinal Chemical Biology Tianjin Key Laboratory of Biosensing and Molecular Recognition Frontiers Science Centre For New Organic Matter Research Centre for Analytical Sciences College of Chemistry School of Medicine and Frontiers Science Center for Cell Responses Nankai University Tianjin P.R. China
Yi‐Fan Wang
State Key Laboratory of Medicinal Chemical Biology Tianjin Key Laboratory of Biosensing and Molecular Recognition Frontiers Science Centre For New Organic Matter Research Centre for Analytical Sciences College of Chemistry School of Medicine and Frontiers Science Center for Cell Responses Nankai University Tianjin P.R. China
Ming‐Yu Zhang
State Key Laboratory of Medicinal Chemical Biology Tianjin Key Laboratory of Biosensing and Molecular Recognition Frontiers Science Centre For New Organic Matter Research Centre for Analytical Sciences College of Chemistry School of Medicine and Frontiers Science Center for Cell Responses Nankai University Tianjin P.R. China
Lei Du
State Key Laboratory of Microbial Technology
Yu‐Xuan Hu
State Key Laboratory of Medicinal Chemical Biology Tianjin Key Laboratory of Biosensing and Molecular Recognition Frontiers Science Centre For New Organic Matter Research Centre for Analytical Sciences College of Chemistry School of Medicine and Frontiers Science Center for Cell Responses Nankai University Tianjin P.R. China
Meng‐Qian Zhang
State Key Laboratory of Medicinal Chemical Biology Tianjin Key Laboratory of Biosensing and Molecular Recognition Frontiers Science Centre For New Organic Matter Research Centre for Analytical Sciences College of Chemistry School of Medicine and Frontiers Science Center for Cell Responses Nankai University Tianjin P.R. China
Shu‐Lin Liu
State Key Laboratory of Medicinal Chemical Biology Tianjin Key Laboratory of Biosensing and Molecular Recognition Frontiers Science Centre For New Organic Matter Research Centre for Analytical Sciences College of Chemistry School of Medicine and Frontiers Science Center for Cell Responses Nankai University Tianjin P.R. China
Chunsheng Wang
Dai‐Wen Pang
Frontiers Science Center for New Organic Matter Research Center for Analytical Sciences College of Chemistry State Key Laboratory of Medicinal Chemical Biology Tianjin Key Laboratory of Biosensing and Molecular Recognition College of Life Sciences Frontiers Science Center for Cell Responses National Demonstration Center for Experimental Chemistry Education Nankai University Tianjin P. R. China