Cell cycle profiling by image and flow cytometry: The optimised protocol for the detection of replicational activity using 5- Bromo-20-deoxyuridine, low concentration of hydrochloric acid and exonuclease III
The result's identifiers
Result code in IS VaVaI
<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F61989592%3A15110%2F17%3A73517698" target="_blank" >RIV/61989592:15110/17:73517698 - isvavai.cz</a>
Result on the web
<a href="http://journals.plos.org/plosone/article/file?id=10.1371/journal.pone.0175880&type=printable" target="_blank" >http://journals.plos.org/plosone/article/file?id=10.1371/journal.pone.0175880&type=printable</a>
DOI - Digital Object Identifier
<a href="http://dx.doi.org/10.1371/journal.pone.0175880" target="_blank" >10.1371/journal.pone.0175880</a>
Alternative languages
Result language
angličtina
Original language name
Cell cycle profiling by image and flow cytometry: The optimised protocol for the detection of replicational activity using 5- Bromo-20-deoxyuridine, low concentration of hydrochloric acid and exonuclease III
Original language description
The approach for the detection of replicational activity in cells using 5-bromo-20-deoxyuridine, a low concentration of hydrochloric acid and exonuclease III is presented in the study. The described method was optimised with the aim to provide a fast and robust tool for the detection of DNA synthesis with minimal impact on the cellular structures using image and flow cytometry. The approach is based on the introduction of breaks into the DNA by the low concentration of hydrochloric acid followed by the subsequent enzymatic extension of these breaks using exonuclease III. Our data showed that the method has only a minimal effect on the tested protein localisations and is applicable both for formaldehyde- and ethanol-fixed cells. The approach partially also preserves the fluorescence of the fluorescent proteins in the HeLa cells expressing Fluorescent Ubiquitin Cell Cycle Indicator. In the case of the short labelling pulses that disabled the use of 5-ethynyl-20-deoxyuridine because of the low specific signal, the described method provided a bright signal enabling reliable recognition of replicating cells. The optimized protocol was also successfully tested for the detection of trifluridine, the nucleoside used as an antiviral drug and in combination with tipiracil also for the treatment of some types of cancer.
Czech name
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Czech description
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Classification
Type
J<sub>imp</sub> - Article in a specialist periodical, which is included in the Web of Science database
CEP classification
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OECD FORD branch
30101 - Human genetics
Result continuities
Project
Result was created during the realization of more than one project. More information in the Projects tab.
Continuities
P - Projekt vyzkumu a vyvoje financovany z verejnych zdroju (s odkazem do CEP)
Others
Publication year
2017
Confidentiality
S - Úplné a pravdivé údaje o projektu nepodléhají ochraně podle zvláštních právních předpisů
Data specific for result type
Name of the periodical
PLoS One
ISSN
1932-6203
e-ISSN
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Volume of the periodical
12
Issue of the periodical within the volume
4
Country of publishing house
US - UNITED STATES
Number of pages
18
Pages from-to
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UT code for WoS article
000399875900063
EID of the result in the Scopus database
2-s2.0-85018511752