title: A sensitive and quantitative autolysosome probe for detecting autophagic activity in live and prestained fixed cells.
abstract: Autophagy is a complex, multi-step and biologically important pathway mediated by autophagosomes and autolysosomes. Accurately dissecting and detecting different stages of autophagy is important to elucidate its molecular mechanism and thereby facilitate the discovery of pharmaceutical molecules. We herein reported a small-molecule synthetic probe, Zn-G 4, which is only fluorescent upon starvation- or chemical agent-induced autophagy within the autolysosome or possible the late endosome/lysosome networks. The probe can be detected by one-photon microscopy, which gives a high signal-to-noise ratio readout of autophagic activity. The pH gradient-independent fluorescence can be detected both in live and prestained fixed cells. Moreover, the fluorescent recording can be used to quantify autophagic activity at a single point without transfection or false positive signals due to protein aggregation. Furthermore, autophagy-induced fluorescence in autolysosomes can also be detected by two-photon microscopy, suggesting potential applications in deep tissue and in vivo. In conclusion, we have developed a sensitive and specific autolysosomal probe that can be used for monitoring autophagy during later stages along with quantitative assays together with widely used early markers or microtubule-associated protein 1 light chain 3 (LC3)-based probes.
authors.affiliation: Beijing National Laboratory for Molecular Sciences, State Key Laboratory of Rare Earth Materials Chemistry and Applications, College of Chemistry and Molecular Engineering, Peking University, Beijing, China.
authors.forename: Juan-Juan
authors.initials: JJ
authors.lastname: Chen
authors.forename: Jing
authors.initials: J
authors.lastname: Jing
authors.forename: Hao
authors.initials: H
authors.lastname: Chang
authors.forename: Yueguang
authors.initials: Y
authors.lastname: Rong
authors.forename: Yang
authors.initials: Y
authors.lastname: Hai
authors.forename: Juan
authors.initials: J
authors.lastname: Tang
authors.forename: Jun-Long
authors.initials: JL
authors.lastname: Zhang
authors.forename: Pingyong
authors.initials: P
authors.lastname: Xu
mesh_terms: D000818:Animals; D001343:Autophagy; D015415:Biomarkers / Q000378:metabolism; D002470:Cell Survival; D004121:Dimethyl Sulfoxide; D005456:Fluorescent Dyes / Q000032:analysis / Q000378:metabolism*; D057809:HEK293 Cells; D006367:HeLa Cells; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008027:Light; D008247:Lysosomes / Q000378:metabolism*; D051379:Mice; D036641:Microscopy, Fluorescence, Multiphoton; D015336:Molecular Probe Techniques; D015335:Molecular Probes / Q000032:analysis / Q000378:metabolism*; D055585:Physical Phenomena; D012997:Solvents; D013194:Staining and Labeling; D013347:Subcellular Fractions / Q000378:metabolism
chemical_list: D015415:Biomarkers; D005456:Fluorescent Dyes; D015335:Molecular Probes; D012997:Solvents; D004121:Dimethyl Sulfoxide
keywords: ZnSalen; autolysosome; autophagy; optical imaging; probe