dc.contributor.author |
Darzynkiewicz, Z |
en |
dc.contributor.author |
Traganos, F |
en |
dc.contributor.author |
Zhao, H |
en |
dc.contributor.author |
Halicka, HD |
en |
dc.contributor.author |
Skommer, Joanna |
en |
dc.contributor.author |
Wlodkowic, Donald |
en |
dc.coverage.spatial |
United States |
en |
dc.date.accessioned |
2011-12-01T23:29:58Z |
en |
dc.date.issued |
2011 |
en |
dc.identifier.citation |
Methods in Cell Biology 103:115-147 2011 |
en |
dc.identifier.issn |
0091-679X |
en |
dc.identifier.uri |
http://hdl.handle.net/2292/9700 |
en |
dc.description.abstract |
This chapter describes molecular mechanisms of DNA damage response (DDR) and presents flow- and image-assisted cytometric approaches to assess these mechanisms and measure the extent of DDR in individual cells. DNA damage was induced by cell treatment with oxidizing agents, UV light, DNA topoisomerase I or II inhibitors, cisplatin, tobacco smoke, and by exogenous and endogenous oxidants. Chromatin relaxation (decondensation) is an early event of DDR chromatin that involves modification of high mobility group proteins (HMGs) and histone H1 and was detected by cytometry by analysis of the susceptibility of DNA in situ to denaturation using the metachromatic fluorochrome acridine orange. Translocation of the MRN complex consisting of Meiotic Recombination 11 Homolog A (Mre11), Rad50 homolog, and Nijmegen Breakage Syndrome 1 (NMR1) into DNA damage sites was assessed by laser scanning cytometry as the increase in the intensity of maximal pixel as well as integral value of Mre11 immunofluorescence. Examples of cytometric detection of activation of Ataxia telangiectasia mutated (ATM), and Check 2 (Chk2) protein kinases using phospho-specific Abs targeting Ser1981 and Thr68 of these proteins, respectively are also presented. We also discuss approaches to correlate activation of ATM and Chk2 with phosphorylation of p53 on Ser15 and histone H2AX on Ser139 as well as with cell cycle position and DNA replication. The capability of laser scanning cytometry to quantify individual foci of phosphorylated H2AX and/or ATM that provides more dependable assessment of the presence of DNA double-strand breaks is outlined. The new microfluidic Lab-on-a-Chip platforms for interrogation of individual cells offer a novel approach for DDR cytometric analysis. |
en |
dc.language |
eng |
en |
dc.publisher |
Elsevier Inc. |
en |
dc.relation.ispartofseries |
Methods in Cell Biology |
en |
dc.rights |
Items in ResearchSpace are protected by copyright, with all rights reserved, unless otherwise indicated. Previously published items are made available in accordance with the copyright policy of the publisher. Details obtained from
http://www.sherpa.ac.uk/romeo/issn/0091-679X/ |
en |
dc.rights.uri |
https://researchspace.auckland.ac.nz/docs/uoa-docs/rights.htm |
en |
dc.subject |
Carcinogens |
en |
dc.subject |
Cell Cycle |
en |
dc.subject |
Cell Cycle Proteins |
en |
dc.subject |
Cell Line, Tumor |
en |
dc.subject |
Chromatin |
en |
dc.subject |
Chromatin Assembly and Disassembly |
en |
dc.subject |
DNA |
en |
dc.subject |
DNA Breaks, Double-Stranded |
en |
dc.subject |
DNA Topoisomerases, Type I |
en |
dc.subject |
Flow Cytometry |
en |
dc.subject |
Histones |
en |
dc.subject |
Humans |
en |
dc.subject |
Image Cytometry |
en |
dc.subject |
Immunohistochemistry |
en |
dc.subject |
Laser Scanning Cytometry |
en |
dc.subject |
Microfluidic Analytical Techniques |
en |
dc.subject |
Neoplasms |
en |
dc.subject |
Oxidation-Reduction |
en |
dc.subject |
Phosphorylation |
en |
dc.subject |
Protein-Serine-Threonine Kinases |
en |
dc.subject |
Radiation, Ionizing |
en |
dc.subject |
Topoisomerase I Inhibitors |
en |
dc.subject |
Tumor Suppressor Proteins |
en |
dc.title |
Analysis of individual molecular events of DNA damage response by flow- and image-assisted cytometry. |
en |
dc.type |
Journal Article |
en |
dc.identifier.doi |
10.1016/B978-0-12-385493-3.00006-1 |
en |
pubs.begin-page |
115 |
en |
pubs.volume |
103 |
en |
dc.rights.holder |
Copyright: 2011 Elsevier Inc. |
en |
dc.identifier.pmid |
21722802 |
en |
pubs.end-page |
147 |
en |
dc.rights.accessrights |
http://purl.org/eprint/accessRights/RestrictedAccess |
en |
pubs.subtype |
Review |
en |
pubs.elements-id |
230072 |
en |
dc.identifier.pii |
B978-0-12-385493-3.00006-1 |
en |
pubs.record-created-at-source-date |
2011-12-02 |
en |
pubs.dimensions-id |
21722802 |
en |