One-step rapid tracking and isolation of senescent cells in cellular systems, tissues, or animal models via GLF16

Sophia Magkouta; Dimitris Veroutis; Athanasios Pousias; Angelos Papaspyropoulos; Kety Giannetti; Natassa Pippa; Nikolaos Lougiakis; Konstantinos Kambas; Nefeli Lagopati; Aikaterini Polyzou; Maria Georgiou; Maria Chountoulesi; Stergios Pispas; Spyros Foutadakis; Efthymios Kyrodimos; Nicole Pouli; Panagiotis Marakos; Athanassios Kotsinas; Panayotis Verginis; Dimitrios Valakos; Giannis Vatsellas; Russell Petty; Dimitris Thanos; Marco Demaria; Konstantinos Evangelou; Raffaella Di Micco; Vassilis G Gorgoulis

doi:10.1016/j.xpro.2024.102929

One-step rapid tracking and isolation of senescent cells in cellular systems, tissues, or animal models via GLF16

STAR Protoc. 2024 Mar 15;5(1):102929. doi: 10.1016/j.xpro.2024.102929. Epub 2024 Mar 8.

Authors

Sophia Magkouta¹, Dimitris Veroutis², Athanasios Pousias³, Angelos Papaspyropoulos², Kety Giannetti⁴, Natassa Pippa⁵, Nikolaos Lougiakis³, Konstantinos Kambas⁶, Nefeli Lagopati⁷, Aikaterini Polyzou⁸, Maria Georgiou³, Maria Chountoulesi⁹, Stergios Pispas¹⁰, Spyros Foutadakis¹¹, Efthymios Kyrodimos¹², Nicole Pouli³, Panagiotis Marakos³, Athanassios Kotsinas⁸, Panayotis Verginis¹³, Dimitrios Valakos¹¹, Giannis Vatsellas¹⁴, Russell Petty¹⁵, Dimitris Thanos¹⁶, Marco Demaria¹⁷, Konstantinos Evangelou⁸, Raffaella Di Micco⁴, Vassilis G Gorgoulis¹⁸

Affiliations

¹ Molecular Carcinogenesis Group, Department of Histology and Embryology, Medical School, National and Kapodistrian University of Athens, 11527 Athens, Greece; Marianthi Simou and G.P. Livanos Labs, 1st Department of Critical Care and Pulmonary Services, School of Medicine, National & Kapodistrian University of Athens, ''Evangelismos'' Hospital, 10676 Athens, Greece.
² Molecular Carcinogenesis Group, Department of Histology and Embryology, Medical School, National and Kapodistrian University of Athens, 11527 Athens, Greece; Biomedical Research Foundation, Academy of Athens, 11527 Athens, Greece.
³ Department of Pharmacy, Division of Pharmaceutical Chemistry, National and Kapodistrian University of Athens, Panepistimiopolis, Zografou, 15771 Athens, Greece.
⁴ San Raffaele Telethon Institute for Gene Therapy (SR-TIGET), IRCCS San Raffaele Scientific Institute, 20132 Milan, Italy.
⁵ Section of Pharmaceutical Technology, Department of Pharmacy, School of Health Sciences, National and Kapodistrian University of Athens, Panepistimioupolis Zografou, 15771 Athens, Greece; Theoretical and Physical Chemistry Institute, National Hellenic Research Foundation, 11635 Athens, Greece.
⁶ Hellenic Pasteur Institute, 11521 Athens, Greece.
⁷ Molecular Carcinogenesis Group, Department of Histology and Embryology, Medical School, National and Kapodistrian University of Athens, 11527 Athens, Greece; Biomedical Research Foundation, Academy of Athens, 11527 Athens, Greece; Laboratory of Biology, Department of Basic Medical Sciences, Medical School, National and Kapodistrian University of Athens, 11527 Athens, Greece.
⁸ Molecular Carcinogenesis Group, Department of Histology and Embryology, Medical School, National and Kapodistrian University of Athens, 11527 Athens, Greece.
⁹ Section of Pharmaceutical Technology, Department of Pharmacy, School of Health Sciences, National and Kapodistrian University of Athens, Panepistimioupolis Zografou, 15771 Athens, Greece.
¹⁰ Theoretical and Physical Chemistry Institute, National Hellenic Research Foundation, 11635 Athens, Greece.
¹¹ Biomedical Research Foundation, Academy of Athens, 11527 Athens, Greece.
¹² First ENT Department, Hippocration Hospital, National Kapodistrian University of Athens, 11527 Athens, GR, Greece.
¹³ Biomedical Research Foundation, Academy of Athens, 11527 Athens, Greece; Laboratory of Immune Regulation and Tolerance, Division of Basic Sciences, University of Crete Medical School, 70013 Heraklion, Greece; Institute of Molecular Biology and Biotechnology, Foundation for Research and Technology, 70013 Heraklion, Greece.
¹⁴ Greek Genome Center, Biomedical Research Foundation, Academy of Athens, 11527 Athens, Greece.
¹⁵ Ninewells Hospital and Medical School, University of Dundee, Dundee DD19SY, UK.
¹⁶ Biomedical Research Foundation, Academy of Athens, 11527 Athens, Greece; Greek Genome Center, Biomedical Research Foundation, Academy of Athens, 11527 Athens, Greece.
¹⁷ European Research Institute for the Biology of Ageing (ERIBA), University Medical Center Groningen, Groningen 9713 AV, The Netherlands.
¹⁸ Molecular Carcinogenesis Group, Department of Histology and Embryology, Medical School, National and Kapodistrian University of Athens, 11527 Athens, Greece; Biomedical Research Foundation, Academy of Athens, 11527 Athens, Greece; Ninewells Hospital and Medical School, University of Dundee, Dundee DD19SY, UK; Faculty Institute for Cancer Sciences, Manchester Academic Health Sciences Centre, University of Manchester, Manchester M20 4GJ, UK; Faculty of Health and Medical Sciences, University of Surrey, Surrey GU2 7YH, UK. Electronic address: vgorg@med.uoa.gr.

Abstract

Identification and isolation of senescent cells is challenging, rendering their detailed analysis an unmet need. We describe a precise one-step protocol to fluorescently label senescent cells, for flow cytometry and fluorescence microscopy, implementing a fluorophore-conjugated Sudan Black-B analog, GLF16. Also, a micelle-based approach allows identification of senescent cells in vivo and in vitro, enabling live-cell sorting for downstream analyses and live in vivo tracking. Our protocols are applicable to cellular systems, tissues, or animal models where senescence is present. For complete details on the use and execution of this protocol, please refer to Magkouta et al.¹.

Keywords: Cancer; Cell Biology; Cell isolation; Cell-based Assays; Flow Cytometry; Molecular/Chemical Probes.

MeSH terms

Animals
Cell Separation
Cellular Senescence*
Flow Cytometry
Fluorescent Dyes*
Models, Animal

Substances

Fluorescent Dyes