How to Identify Senescence in Cells and Tissue | CST Tech Tips

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Senescence is a stress-inducible and irreversible form of cell cycle arrest. This video briefly reviews the features of cellular senescence and introduces a three-stage workflow for identifying senescent cells in your samples using markers. We also share some practical tips to help you avoid formation of blue crystals when using X-gal to stain for β-galactosidase.

0:00 Intro
0:21 What defines cellular senescence?
0:46 The Hayflick limit
1:45 Cellular hallmarks of senescence
2:50 Screening for senescence
3:21 β-galactosidase staining tips
4:17 Verification with additional biomarkers
4:37 Specific types of senescence and SASP
5:41 Quantitative readouts for senescence


🔹 Download the Cellular Senescence handbook: https://cst-science.com/fqcqwy
🔹 Senescence β-Galactosidase Staining Kit (Fluorescence, Flow Cytometry) #35302: https://cst-science.com/i9ykac
🔹 Senescence β-Galactosidase Staining Kit Kit (Fluorescence, Plate-Based) #23833: https://cst-science.com/ycj142
🔹 Senescence β-Galactosidase Staining Kit #9860: https://cst-science.com/62klcj
🛒 More kits and markers for identifying sensescence: https://cst-science.com/2fceuo
🖱 Interactive Senescence Signaling Pathway Diagram: https://cst-science.com/s0wvm9

Partial transcript:
What is cellular senescence, and how can I identify it in my samples? I’m Srikanth, Senior Product Scientist at Cell Signaling Technology, and this is CST Tech Tips.

If your research is related to development, aging, or diseases such as cancer, you may be interested in investigating whether senescence is involved in your model system. But what defines cellular senescence?

First, let’s note that the term senescence may be used broadly to refer to the aging of organisms, while cellular senescence, also referred to as replicative senescence, focuses on the cellular scale. Senescent cells withdraw from the cell cycle and stop dividing, but they stay metabolically active. This is a normal feature of both aging and development in vivo, and it can be observed in vitro as well.

You probably know that when propagating cell lines, a best practice is to mark the passage number on the culture flask or dish, and carefully track the number of times cells have been split. There are several reasons to do this, one of which is that if you continue splitting non-transformed cells long enough, you’ll eventually observe that they slow and then stop dividing. This finite limit of the number of times cell populations will divide before becoming senescent is known as the Hayflick Limit, reported in a 1961 paper by Hayflick and Moorehead.

Senescence normally occurs during development and aging, but it can also be triggered by the DNA damage response and stress signaling pathways, and it has been linked to various age-related pathologies including cancers and cardiovascular, metabolic, and neurodegenerative diseases.

Researchers working in these fields often rely on senescence biomarkers to distinguish it from quiescence, which allows re-entry into the cell cycle, and from terminal differentiation, which involves cell cycle exit as the result of normal developmental cues and programmed differentiation.

Cellular senescence is associated with a flattened cell morphology, increased secretory and pro-inflammatory activity, chromatin reorganization, and shortening of telomeres. We’ll highlight a few specific senescence markers in this video.

But before we do, I want to emphasize that there is no universal marker and no single assay to determine senescence. For example, not all senescent cells will be positive for every biomarker, and some of these biomarkers may also be observed in non-senescent phenomena such as quiescence or apoptosis.

However, by examining multiple biomarkers in combination, you can increase your confidence in your results. A three-stage workflow, starting with screening, followed by co-staining for verification, and then moving on to analysis of markers for specific types of senescence, is recommended.

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