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Detection of apoptosis by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling and acridine orange in Drosophila embryos and adult male gonads

Nature Protocols volume 1, pages 1725–1731 (2006)Cite this article

Abstract

In Drosophila, vast numbers of cells undergo apoptosis during normal development. In addition, excessive apoptosis can be induced in response to a variety of stress or injury paradigms, including DNA damage, oxidative stress, nutrient deprivation, unfolded proteins and mechanical tissue damage. Two of the most commonly used methods to label apoptotic cells in Drosophila are terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) for fixed tissues and acridine orange (AO) staining for live embryos or tissues. Here, we describe protocols for labeling apoptotic cells in Drosophila embryos and adult male gonads. Slightly modified protocols can also be applied for other Drosophila tissues. The AO protocol is quick, simple and allows real-time imaging of doomed cells in live tissues. However, it is difficult to combine with conventional counterstains or Ab labeling. On the other hand, this functionality is readily afforded by the TUNEL protocol, which permits the detection of apoptotic cells in fixed tissues. These staining procedures can be completed in 1–2 d.

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Figure 1: TUNEL staining of Drosophila testes.
Figure 2: AO staining of embryos (a,b) and testes (c,d).
Figure 3: Dissecting male gonads.

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Acknowledgements

We thank C. Gafuik and G. Rieckhof for critically reading the manuscript and M. Bader for help with Figure 3. E.A. was a fellow of the Charles H. Revson Foundation and H.S. is an investigator with the Howard Hughes Medical Institute. Part of this work was supported by NIH grant RO1 GM60124 to H.S.

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  1. Eli Arama

    Present address: Department of Molecular Genetics, Weizmann Institute of Science, Rehovot, Israel

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  1. Howard Hughes Medical Institute, Strang Laboratory of Cancer Research, The Rockefeller University, 1230 York Avenue, New York, 10021, New York, USA

    Eli Arama & Hermann Steller

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Correspondence to Hermann Steller.

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Arama, E., Steller, H. Detection of apoptosis by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling and acridine orange in Drosophila embryos and adult male gonads. Nat Protoc 1, 1725–1731 (2006). https://doi.org/10.1038/nprot.2006.235

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