ArticleMethods in molecular biology (Clifton, N.J.)2011
Detection of DNA strand breaks in apoptotic cells by flow- and image-cytometry.
Article in Methods in molecular biology (Clifton, N.J.), 2011. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 10 papers.
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Who cites it
10 citing papers in PubMed, 19 citations in OpenAlex.
- DNA Double Strand Break and Response Fluorescent Assays: Choices and Interpretation.International journal of molecular sciences · 2024Review
- A Light in the Dark: Uncovering Wolbachia-Host Interactions Using Fluorescence Imaging.Methods in molecular biology (Clifton, N.J.) · 2024Article
- The AMPK activator metformin improves recovery from demyelination by shifting oligodendrocyte bioenergetics and accelerating OPC differentiation.Frontiers in cellular neuroscience · 2023Article
- Synthesis and Anticancer Activity of Indole-Functionalized Derivatives of Betulin.Pharmaceutics · 2022Article
- Mechanical competition triggered by innate immune signaling drives the collective extrusion of bacterially infected epithelial cells.Developmental cell · 2021Article
- Mechanical Regulation of Apoptosis in the Cardiovascular System.Annals of biomedical engineering · 2021Review
- The Calotropis Gigantea Methanolic Extract Induces Apoptosis in Human Breast Carcinoma Cells.Iranian journal of medical sciences · 2019Article
- An overview of apoptosis assays detecting DNA fragmentation.Molecular biology reports · 2018Review
- Article
- TUNEL labeling with BrdUTP/anti-BrdUTP greatly underestimates the level of sperm DNA fragmentation in semen evaluation.PloS one · 2017Article
Corrections and comments
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Authors and funding
2 authors at 1 institution in 1 country.
Funding
Abstract
Extensive DNA fragmentation that generates a multitude of DNA double-strand breaks (DSBs) is a hallmark of apoptosis. A widely used approach to identify apoptotic cells relies on labeling DSBs in situ with fluorochromes. Flow or image cytometry is then used to detect and quantify apoptotic cells labeled this way. We developed several variants of the methodology that is based on the use of exogenous terminal deoxynucleotidyl transferase (TdT) to label 3'-OH ends of the DSBs with fluorochromes, defined as the TUNEL assay. This chapter describes the variant based on DSBs labeling using 5-Bromo-2'-deoxyuridine-5'-triphosphate (BrdUTP) as a TdT substrate and the incorporated BrdU is subsequently detected immunocytochemically with anti-BrdU antibody. We also describe modifications of the protocol that allow using other than BrdUTP deoxyribonucleotides to label DSBs. Concurrent differential staining of cellular DNA and multiparameter analysis of cells by flow- or image cytometry enables one to correlate the induction of apoptosis with the cell cycle phase. Examples of the detection of apoptotic cells in cultures of human leukemic cell lines treated with TNF-α and DNA topoisomerase I inhibitor topotecan are presented. The protocol can be applied to the cells growing in vitro, treated ex vivo with cytotoxic drugs as well as to clinical samples.
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