In situ DNA damaging foci analysis on metaphase chromosomes

Abstract

DNA damage foci such as ionizing radiation-induced foci (IRIF) can visually distinguish the location and number of specific types of DNA damages. This method is widely used to detect DNA damage in interphase cells. These DNA damage foci can be also visualized on metaphase chromosomes. The technique has an advantage as it provides an easy method of quantifying chromosomal DNA damage. Radiation-induced DNA double strand breaks can be assessed for gamma-H2AX foci formation on metaphase chromosomes. Gamma-H2AX foci can be observed at the break point of chromosomes and can persist in newly repair chromosomes. Foci observation may be advantageous compared to classical cytogenetic analysis due to less time required for analysis. Metaphase DNA damage analysis can be also used for the estimation of DNA damage persistence in daughter cells and capacity of DNA repair. Not only DNA double strand breaks can be visualized, but also other types of DNA damage and modification such as oxidative damage, crosslinking, and methylation of DNA can be visualized with appropriate antibodies. This IRIF immunostaining technique can be combined with FISH analysis for the immunoFISH method.