title: Measurement of apoptosis by SCAN©, a system for counting and analysis of fluorescently labelled nuclei.
abstract: Apoptosis-like programmed cell death (A-PCD) is a universal process common to all types of eukaryotic organisms. Because A-PCD-associated processes are conserved, it is possible to define A-PCD by a standard set of markers. Many of the popular methods to measure A-PCD make use of fluorescent ligands that change in intensity or cellular localization during A-PCD. In single cell organisms, it is possible to quantify levels of A-PCD by scoring the number of apoptotic cells using flow cytometry instruments. In a multicellular organism, quantification of A-PCD is more problematic due to the complex nature of the tissue. The situation is further complicated in filamentous fungi, in which nuclei are divided between compartments, each containing a number of nuclei, which can also migrate between the compartments. We developed SCAN©, a System for Counting and Analysis of Nuclei, and used it to measure A-PCD according to two markers - chromatin condensation and DNA strand breaks. The package includes three modules designed for counting the number of nuclei in multi-nucleated domains, scoring the relative number of nuclei with condensed chromatin, and calculating the relative number of nuclei with DNA strand breaks. The method provides equal or better results compared with manual counting, the analysis is fast and can be applied on large data sets. While we demonstrated the utility of the software for measurement of A-PCD in fungi, the method is readily adopted for measurement of A-PCD in other types of multicellular specimens.
authors.affiliation: Department of Molecular Biology and Ecology of Plants, Faculty of Life Sciences, Tel Aviv University, Tel Aviv 69978, Israel.
authors.forename: Neta
authors.initials: N
authors.lastname: Shlezinger
authors.affiliation: Department of Molecular Biology and Ecology of Plants, Faculty of Life Sciences, Tel Aviv University, Tel Aviv 69978, Israel. ; Department of Physical Electronics, Fleischman Faculty of Engineering, Tel Aviv University, Tel Aviv 69978, Israel.
authors.forename: Elad
authors.initials: E
authors.lastname: Eizner
authors.affiliation: Department of Physical Electronics, Fleischman Faculty of Engineering, Tel Aviv University, Tel Aviv 69978, Israel.
authors.forename: Stas
authors.initials: S
authors.lastname: Dubinchik
authors.affiliation: Department of Molecular Biology and Ecology of Plants, Faculty of Life Sciences, Tel Aviv University, Tel Aviv 69978, Israel.
authors.forename: Anna
authors.initials: A
authors.lastname: Minz-Dub
authors.affiliation: Department of Molecular Biology and Ecology of Plants, Faculty of Life Sciences, Tel Aviv University, Tel Aviv 69978, Israel.
authors.forename: Rachel
authors.initials: R
authors.lastname: Tetroashvili
authors.affiliation: Department of Molecular Biology and Ecology of Plants, Faculty of Life Sciences, Tel Aviv University, Tel Aviv 69978, Israel.
authors.forename: Adi
authors.initials: A
authors.lastname: Reider
authors.affiliation: Department of Molecular Biology and Ecology of Plants, Faculty of Life Sciences, Tel Aviv University, Tel Aviv 69978, Israel.
authors.forename: Amir
authors.initials: A
authors.lastname: Sharon
keywords: DNA condensation; PCD; TUNEL; apoptosis; fungi