Senescence detection using reflected light.
SABG
X‐gal
adipocytes
beta‐galactosidase
brightfield
reflected light
senescence
western blot
Journal
Aging cell
ISSN: 1474-9726
Titre abrégé: Aging Cell
Pays: England
ID NLM: 101130839
Informations de publication
Date de publication:
05 Aug 2024
05 Aug 2024
Historique:
revised:
10
07
2024
received:
26
02
2024
accepted:
18
07
2024
medline:
6
8
2024
pubmed:
6
8
2024
entrez:
5
8
2024
Statut:
aheadofprint
Résumé
Senescence is an important cellular program occurring in development, tissue repair, cancer, and aging. Increased senescence is also associated with disease states, including obesity and Type 2 diabetes (T2D). Characterizing and quantifying senescent cells at a single cell level has been challenging and particularly difficult in large primary cells, such as human adipocytes. In this study, we present a novel approach that utilizes reflected light for accurate senescence-associated beta-galactosidase (SABG) staining measurements, which can be integrated with immunofluorescence and is compatible with primary mature adipocytes from both human and mouse, as well as with differentiated 3T3-L1 cells. This technique provides a more comprehensive classification of a cell's senescent state by incorporating multiple criteria, including robust sample-specific pH controls. By leveraging the precision of confocal microscopy to detect X-gal crystals using reflected light, we achieved superior sensitivity over traditional brightfield techniques. This strategy allows for the capture of all X-gal precipitates in SABG-stained samples, revealing diverse X-gal staining patterns and improved detection sensitivity. Additionally, we demonstrate that reflected light outperforms western blot analysis for the detection and quantification of senescence in mature human adipocytes, as it offers a more accurate representation of SABG activity. This detection strategy enables a more thorough investigation of senescent cell characteristics and specifically a deeper look at the relationship between adipocyte senescence and obesity associated disorders, such as T2D.
Types de publication
Journal Article
Langues
eng
Sous-ensembles de citation
IM
Pagination
e14295Subventions
Organisme : Vetenskapsrådet
ID : 2022-01236
Organisme : Knut och Alice Wallenbergs Stiftelse
ID : 2020.0118
Organisme : Cancerfonden
ID : 22 2420
Organisme : Vallee Foundation
ID : C5471234
Organisme : Karolinska Institutet
ID : C5471162
Organisme : Karolinska Institutet
ID : C5472022
Organisme : Mark Foundation For Cancer Research
ID : C5477023
Organisme : Novo Nordisk Fonden
ID : C5475033
Organisme : Novo Nordisk Fonden
ID : NNF21OC0070149
Organisme : The Erling-Persson Foundation
ID : 140609
Informations de copyright
© 2024 The Author(s). Aging Cell published by Anatomical Society and John Wiley & Sons Ltd.
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