Genotoxicity and DNA Damage and Repair
Genotoxicity and DNA Damage and Repair
Last Updated on Wednesday, 23 November 2011 15:35
Overview
High-content enables the quantification of genomics instability resulting from DNA damage in the form of single- and double-strand breaks and affects the control of cell cycle checkpoints. Several indicators of these events include phosphorylation of Histone H2AX, ATM kinase, Chk2 or p53, micronucleus formation or the Ku 70/80 activation of DNA protein kinases. Cell cycle and proliferation can also be monitored by BrdU incorporation during DNA replication (S phase), phosphorylation of Histone H3 (M phase), or cell proliferation specific gene expression such as Ki67 (G1, S, G2 and M phase), among others. Micronucleus (MN) formation is a hallmark of genetic toxicity; as such, micronuclei are used as indicators of genotoxicity caused by drug candidates or environmental toxins.The multiparameter nature of high-content allows for the multiplexing of targets and deeper understanding of genomic instability and its repair.
Application Example
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Micronucleus formation may result from clastogens, which cause chromosomal breaks, and/or aneugens, which affect the spindle apparatus and result in the loss of whole chromosomes during anaphase. The in vitro micronucleus assay is among a new set of genetic toxicology assays in which cultured cells are treated and scored for micronucleus induction. This enables screening for indicators of genetic toxicity earlier in the development of therapeutic candidates, which can in turn impact attrition rates and reduce costs. The Thermo Scientific ArrayScan VTI HCS Reader and associated Micronucleus BioApplication allows monitoring of genotoxicity by examining the frequency of micronucleus formation in single cells. Automated measurement of genotoxicity in intact cells represents a more functional and accurate assay compared to the time-consuming method of manually scoring cells. The Micronucleus BioApplication has been optimized for the “cytokinesis-block” method of measuring micronucleus induction. In addition to genotoxicity, it automatically determines both the percentage of cells that contain micronuclei and the ratio of multinucleated to mononucleated cells for monitoring cell-cycle delay, an early indicator of cytotoxicity. Alternatively, the BioApplication can be configured to examine populations of mononucleated cells for micronuclei induction.High-content hence decreases time to quantitatively analyze populations of cells in multiple |
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Assays and Reagents
Thermo Scientific Cellomics HCS Reagents and Assays are available for a wide variety of targets. Each assay offers all the components necessary to perform the assay as well as a validated protocol, with step-by-step instructions for use on any of the Thermo Scientific HCS Readers as well as other instrumetation and standard fluorescence microscopes. Reagents and Assays are available in kit form (everything in the box) or as components where based on your need, so you may purchase all or al-la-carte reagents for the assay. To order components for a legacy kit please visit our ReagentFinder page.
