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Article

Apoptosis Quantification in Tissue: Development of a Semi-Automatic Protocol and Assessment of Critical Steps of Image Processing

1
Laboratoire de Génétique et de Biologie Cellulaire (LGBC), UVSQ, Université Paris-Saclay, 78000 Versailles, France
2
Ecole Pratique des Hautes Etudes, PSL Research University, 75014 Paris, France
*
Authors to whom correspondence should be addressed.
Present adress: Unité Biologie des ARN des Pathogènes Fongiques, Institut Pasteur, F-75015 Paris, France.
Academic Editor: Stéphen T. Manon
Biomolecules 2021, 11(10), 1523; https://doi.org/10.3390/biom11101523
Received: 22 July 2021 / Revised: 12 October 2021 / Accepted: 12 October 2021 / Published: 15 October 2021
(This article belongs to the Special Issue State-of-the-Art Cell Death in France 2020-2021)
Apoptosis is associated with numerous phenotypical characteristics, and is thus studied with many tools. In this study, we compared two broadly used apoptotic assays: TUNEL and staining with an antibody targeting the activated form of an effector caspase. To compare them, we developed a protocol based on commonly used tools such as image filtering, z-projection, and thresholding. Even though it is commonly used in image-processing protocols, thresholding remains a recurring problem. Here, we analyzed the impact of processing parameters and readout choice on the accuracy of apoptotic signal quantification. Our results show that TUNEL is quite robust, even if image processing parameters may not always allow to detect subtle differences of the apoptotic rate. On the contrary, images from anti-cleaved caspase staining are more sensitive to handle and necessitate being processed more carefully. We then developed an open-source Fiji macro automatizing most steps of the image processing and quantification protocol. It is noteworthy that the field of application of this macro is wider than apoptosis and it can be used to treat and quantify other kind of images. View Full-Text
Keywords: apoptosis; TUNEL; caspase; image processing; thresholding; signal quantification; Drosophila apoptosis; TUNEL; caspase; image processing; thresholding; signal quantification; Drosophila
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MDPI and ACS Style

de Noiron, J.; Hoareau, M.; Colin, J.; Guénal, I. Apoptosis Quantification in Tissue: Development of a Semi-Automatic Protocol and Assessment of Critical Steps of Image Processing. Biomolecules 2021, 11, 1523. https://doi.org/10.3390/biom11101523

AMA Style

de Noiron J, Hoareau M, Colin J, Guénal I. Apoptosis Quantification in Tissue: Development of a Semi-Automatic Protocol and Assessment of Critical Steps of Image Processing. Biomolecules. 2021; 11(10):1523. https://doi.org/10.3390/biom11101523

Chicago/Turabian Style

de Noiron, Juliette, Marion Hoareau, Jessie Colin, and Isabelle Guénal. 2021. "Apoptosis Quantification in Tissue: Development of a Semi-Automatic Protocol and Assessment of Critical Steps of Image Processing" Biomolecules 11, no. 10: 1523. https://doi.org/10.3390/biom11101523

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