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A Fully-Automated Senescence Test (FAST) for the high-throughput quantification of senescence-associated markers
Francesco Neri1,2, Selma N Takajjart1, Chad A Lerner1
1Buck Institute for Research on Aging, Novato, CA, USA.
Geroscience
|June 13, 2024
Summary
We developed the Fully-Automated Senescence Test (FAST) to accurately quantify senescent cells using an image-based method. This high-throughput approach overcomes challenges in aging research by providing unbiased, single-cell analysis of senescence markers.
Area of Science:
- Cell Biology
- Aging Research
- Biotechnology
Background:
- Cellular senescence is a key factor in aging and age-related diseases.
- Accurate quantification of senescent cells is difficult due to a lack of specific markers and unbiased methods.
- Existing methods often struggle with high-throughput analysis and can be prone to false positives.
Purpose of the Study:
- To introduce the Fully-Automated Senescence Test (FAST), an image-based, high-throughput method for single-cell senescence assessment.
- To provide an unbiased and user-friendly tool for quantifying senescence burden in cultured cells.
- To enable large-scale experiments and compound screening in aging research.
Main Methods:
- FAST quantifies senescence-associated β-galactosidase activity (SA-β-Gal), proliferation arrest (EdU incorporation), and cell morphology (nuclear area).
- The workflow includes automated microplate image acquisition, processing, and data analysis with both threshold-based and machine learning approaches.
- Standardization was achieved through optical density quantification of SA-β-Gal, background controls, and automated processes.
Main Results:
- FAST accurately quantifies senescence burden, is cell-type agnostic, and robust to different microscope setups.
- The method effectively reduces false-positive staining, a common artifact in senescence assays.
- A modest correlation was observed between X-Gal and C12FDG SA-β-Gal staining, highlighting differences in detection.
- Proof of concept for compound screening demonstrates FAST's utility in identifying senescence-modifying agents.
Conclusions:
- FAST offers a rapid, unbiased, and user-friendly solution for quantifying cellular senescence in culture.
- This method facilitates previously impractical large-scale experiments and compound screening for aging research.
- FAST addresses critical needs in the aging field for robust and high-throughput senescence assessment.
