4.0 Article

Whole-mount Senescence-Associated Beta-Galactosidase (SA-β-GAL) Activity Detection Protocol for Adult Zebrafish

Journal

BIO-PROTOCOL
Volume 12, Issue 13, Pages -

Publisher

BIO-PROTOCOL
DOI: 10.21769/BioProtoc.4457

Keywords

SA-beta-GAL; Senescence; Aging; Whole organism; Zebrafish

Categories

Funding

  1. FCT (Portugal) [LISBOA-01-0145-FEDER-022170]
  2. Lisboa2020, under the PORTUGAL2020 agreement (European Regional Development Fund) [LISBOA-01-0145-FEDER-022170]
  3. Ville de Nice
  4. Fondation pour la Recherche Medicale FRM [EQU201903007804]
  5. Agence Nationale de la Recherche [ANR-21-CE14-0054]

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This study presents a protocol for detecting senescent cell accumulation in adult zebrafish organs using SA-beta-GAL assay and identifies previously unreported organs with increased cellular senescence in telomerase mutants.
Senescence-associated beta-galactosidase (SA-beta-GAL) is an enzyme that accumulates in the lysosomes of senescent cells, where it hydrolyses beta-galactosides. With p16, it represents a well-recognized biomarker used to assess senescence both in vivo and in cell culture. The use of a chromogenic substrate, such as 5 -bromo-4-chloro-3-indoyl-beta-d-galactopyranoside (X-Gal), allows the detection of SA-beta-GAL activity at pH 6.0 by the release of a visible blue product. Senescence occurs during aging and is part of the aging process itself. We have shown that prematurely aged zebrafish accumulate senescent cells detectable by SA-beta-GAL staining in different tissues, including testis and gut. Here, we report a detailed protocol to perform an SA-beta-GAL assay to detect senescent cell accumulation across the entire adult zebrafish organism (Danio rerio). We also identify previously unreported organs that show increased cell senescence in telomerase mutants, including the liver and the spinal cord.

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