title: How to unveil self-quenched fluorophores and subsequently map the subcellular distribution of exogenous peptides.
abstract: Confocal laser scanning microscopy (CLSM) is the most popular technique for mapping the subcellular distribution of a fluorescent molecule and is widely used to investigate the penetration properties of exogenous macromolecules, such as cell-penetrating peptides (CPPs), within cells. Despite the membrane-association propensity of all these CPPs, the signal of the fluorescently labeled CPPs did not colocalize with the plasma membrane. We studied the origin of this fluorescence extinction and the overall consequence on the interpretation of intracellular localizations from CLSM pictures. We demonstrated that this discrepancy originated from fluorescence self-quenching. The fluorescence was unveiled by a "dilution" protocol, i.e. by varying the ratio fluorescent/non-fluorescent CPP. This strategy allowed us to rank with confidence the subcellular distribution of several CPPs, contributing to the elucidation of the penetration mechanism. More generally, this study proposes a broadly applicable and reliable method to study the subcellular distribution of any fluorescently labeled molecules.
authors.affiliation: Sorbonne Universités, UPMC Univ Paris 06, LBM, 4, Place Jussieu, 75005 Paris, France.|Ecole Normale Supérieure - PSL research University, Département de Chimie, 24 Rue Lhomond, 75005 Paris, France.|CNRS, UMR 7203, LBM, Paris, France.
authors.forename: Jean-Marie
authors.initials: JM
authors.lastname: Swiecicki
authors.affiliation: Sorbonne Universités, UPMC Univ Paris 06, LBM, 4, Place Jussieu, 75005 Paris, France.|Ecole Normale Supérieure - PSL research University, Département de Chimie, 24 Rue Lhomond, 75005 Paris, France.|CNRS, UMR 7203, LBM, Paris, France.
authors.forename: Frédéric
authors.initials: F
authors.lastname: Thiebaut
authors.affiliation: Sorbonne Universités, UPMC Univ Paris 06, LBM, 4, Place Jussieu, 75005 Paris, France.|Ecole Normale Supérieure - PSL research University, Département de Chimie, 24 Rue Lhomond, 75005 Paris, France.|CNRS, UMR 7203, LBM, Paris, France.
authors.forename: Margherita
authors.initials: M
authors.lastname: Di Pisa
authors.affiliation: Sorbonne Universités, UPMC Univ Paris 06, PHENIX, 4 Place Jussieu, 75005 Paris, France.|CNRS, UMR 8234, PHENIX, Paris, France.
authors.forename: Simon
authors.initials: S
authors.lastname: Gourdin-Bertin
authors.affiliation: Sorbonne Universités, UPMC Univ Paris 06, LBM, 4, Place Jussieu, 75005 Paris, France.|Ecole Normale Supérieure - PSL research University, Département de Chimie, 24 Rue Lhomond, 75005 Paris, France.|CNRS, UMR 7203, LBM, Paris, France.
authors.forename: Julien
authors.initials: J
authors.lastname: Tailhades
authors.affiliation: Sorbonne Universités, UPMC Univ Paris 06, LBM, 4, Place Jussieu, 75005 Paris, France.|Ecole Normale Supérieure - PSL research University, Département de Chimie, 24 Rue Lhomond, 75005 Paris, France.|CNRS, UMR 7203, LBM, Paris, France.
authors.forename: Christelle
authors.initials: C
authors.lastname: Mansuy
authors.affiliation: Sorbonne Universités, UPMC Univ Paris 06, LBM, 4, Place Jussieu, 75005 Paris, France.|Ecole Normale Supérieure - PSL research University, Département de Chimie, 24 Rue Lhomond, 75005 Paris, France.|CNRS, UMR 7203, LBM, Paris, France.
authors.forename: Fabienne
authors.initials: F
authors.lastname: Burlina
authors.affiliation: Sorbonne Universités, UPMC Univ Paris 06, LBM, 4, Place Jussieu, 75005 Paris, France.|INSERM-ERL 1157, CHU Saint Antoine, 27 rue de Chaligny, 75012 Paris, France.|AP-HP, Hôpital Saint Antoine, 75012 Paris, France.|INRA, UR892, Virologie et Immunologie Moléculaires, 78350 Jouy-en-Jossas, France.
authors.forename: Serge
authors.initials: S
authors.lastname: Chwetzoff
authors.affiliation: Sorbonne Universités, UPMC Univ Paris 06, LBM, 4, Place Jussieu, 75005 Paris, France.|AP-HP, Hôpital Saint Antoine, 75012 Paris, France.|INRA, UR892, Virologie et Immunologie Moléculaires, 78350 Jouy-en-Jossas, France.
authors.forename: Germain
authors.initials: G
authors.lastname: Trugnan
authors.affiliation: Sorbonne Universités, UPMC Univ Paris 06, LBM, 4, Place Jussieu, 75005 Paris, France.|Ecole Normale Supérieure - PSL research University, Département de Chimie, 24 Rue Lhomond, 75005 Paris, France.|CNRS, UMR 7203, LBM, Paris, France.
authors.forename: Gérard
authors.initials: G
authors.lastname: Chassaing
authors.affiliation: Sorbonne Universités, UPMC Univ Paris 06, LBM, 4, Place Jussieu, 75005 Paris, France.|Ecole Normale Supérieure - PSL research University, Département de Chimie, 24 Rue Lhomond, 75005 Paris, France.|CNRS, UMR 7203, LBM, Paris, France.
authors.forename: Solange
authors.initials: S
authors.lastname: Lavielle
mesh_terms: D002463:Cell Membrane Permeability; D057846:Cell-Penetrating Peptides / Q000737:chemistry / Q000378:metabolism*; D005456:Fluorescent Dyes / Q000737:chemistry* / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D007476:Ionophores; D018613:Microscopy, Confocal
chemical_list: D057846:Cell-Penetrating Peptides; D005456:Fluorescent Dyes; D007476:Ionophores
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