Multiparameter flow cytometric detection and quantification of senescent cells in vitro
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RESEARCH ARTICLE
Multiparameter flow cytometric detection and quantification of senescent cells in vitro Adeolu Badi Adewoye Alexandra Stolzing
. Dimitris Tampakis . Antonia Follenzi
.
Received: 28 May 2020 / Accepted: 30 July 2020 Ó The Author(s) 2020
Abstract It has been over half a century since cellular senescence was first noted and characterized, and yet no consensus senescent marker has been reliably established. This challenge is compounded by the complexity and heterogenic phenotypes of senescent cells. This necessitates the use of multiple biomarkers to confidently characterise senescent cells. Despite cytochemical staining of senescence associated-beta-galactosidase being a single marker approach, as well as being time and labour-intensive, it remains the most popular detection method. We have developed an alternative flow cytometry-based method that simultaneously quantifies multiple senescence markers at a single-cell resolution. In this study, Adeolu Badi Adewoye and Dimitris Tampakis have contributed equally to this work.
we applied this assay to the quantification of both replicative and induced senescent primary cells. Using this assay, we were able to quantify the activity level of SA b-galactosidase, the expression level of p16INK4a and cH2AX in these cell populations. Our results show this flow cytometric approach to be sensitive, robust, and consistent in discriminating senescent cells in different cell senescence models. A strong positive correlation between these commonlyused senescence markers was demonstrated. The method described in this paper can easily be scaled up to accommodate high-throughput screening of senescent cells in applications such as therapeutic cell preparation, and in therapy-induced senescence following cancer treatment. Keywords Senescence Mesenchymal stem cells Aging Flow cytometry Quantification
Electronic supplementary material The online version of this article (doi:https://doi.org/10.1007/s10522-020-09893-9) contains supplementary material, which is available to authorized users. A. B. Adewoye D. Tampakis A. Stolzing (&) Centre for Biological Engineering, School of Mechanical, Electrical and Manufacturing Engineering, Loughborough University, Loughborough LE11 3TU, UK e-mail: [email protected] A. B. Adewoye Institute of Inflammation and Ageing, College of Medical and Dental Sciences, University of Birmingham, Birmingham B15 2TT, UK
D. Tampakis Division of Cancer Studies, King’s College London, London, UK A. Follenzi Department of Health Sciences, Universita` del Piemonte Orientale ‘‘A. Avogadro,’’, 28100 Novara, Italy
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Biogerontology
Introduction Emerging evidence suggests that the accumulation of senescent cells in tissues and organs may underlie ageing and age-related diseases (Krishnamurthy et al. 2004; Chen et al. 2007; Yanai and Fraifeld 2018). For instance, accumulation of senescent cells has been reported in ageing primate, rodent, and human tissues (Dimri et al. 1995; Herbig et a
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