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FISH Analysis of Aging-Associated Aneuploidy in Neurons and Nonneuronal Brain Cells

  • Grasiella A. Andriani
  • Cristina MontagnaEmail author
Protocol
Part of the Neuromethods book series (NM, volume 131)

Abstract

Aging is a ubiquitous complex process characterized by tissue degeneration and loss of cellular fitness. Genome instability (GIN) has long been implicated as a main causal factor in aging. The most severe form of genomic instability is whole chromosome instability (W-CIN), a state where dysfunction in chromosome segregation leads to whole chromosomes gains and losses. Aneuploidy is commonly linked to pathological states. It is a hallmark of spontaneous abortions and birth defects and it is observed virtually in every human tumor. There is mounting evidence that W-CIN increases with age, with the underlying hypothesis that some of the age-related loss of fitness phenotypes may be the result of W-CIN. Methodologically, the detection of stochastic W-CIN during the aging process poses unique challenges: aneuploid cells are scattered among diploid cells and, contrary to the cancer genome where aneuploidy is present in the background of massive ploidy changes, the number of aneuploid chromosome per cells is usually low (few per cell). Aging-associated aneuploidy is also largely stochastic or with limited clonal expansion. Therefore analysis at the single-cell level and the examination of a large number of cells is necessary. Here we describe a modification of the standard fluorescent in situ hybridization (FISH) protocol adapted for the detection of low-frequency mosaic aneuploidy in interphase cells isolated from the adult brain or within frozen tissue sections. This approach represents a straightforward method for the single-cell analysis of W-CIN in mammalian cells. It is based on the combination of four probes mapping to two different chromosomes and analysis of interphase cells, highly reducing false positives and enabling studying W-CIN also in post-mitotic tissues.

Key words

FISH (Fluorescent in situ hybridization) Fluorophores Aneuploidy Interphase FISH Genomic instability (GIN) Whole chromosome instability (W-CIN) Aging Brain NeuN+ NeuN− 

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© Springer Science+Business Media LLC 2017

Authors and Affiliations

  1. 1.Department of GeneticsAlbert Einstein College of Medicine, Yeshiva UniversityBronxUSA
  2. 2.Department of PathologyAlbert Einstein College of Medicine, Yeshiva UniversityBronxUSA

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