Quantitation of Apurinic/Apyrimidinic Sites in Isolated DNA and in Mammalian Tissue with a Reduced Level of Artifacts.
Journal
Analytical chemistry
ISSN: 1520-6882
Titre abrégé: Anal Chem
Pays: United States
ID NLM: 0370536
Informations de publication
Date de publication:
04 06 2019
04 06 2019
Historique:
pubmed:
6
5
2019
medline:
10
9
2020
entrez:
7
5
2019
Statut:
ppublish
Résumé
The apurinic/apyrimidinic (AP) site is a common lesion of DNA damage. The levels of AP sites reported in the literature cover a wide range, which is primarily due to the artifactual generation or loss of AP sites during processing of the DNA. Herein, we have developed a method for quantitating AP sites with a largely reduced level of artifacts by derivatizing AP sites before DNA isolation. A rapid digestion of nuclear protein was performed to minimize enzymatic DNA repair, followed by direct derivatization of AP sites in the nuclear lysate with O-(pyridin-3-yl-methyl)hydroxylamine, yielding an oxime derivative that is stable through the subsequent DNA processing steps. Quantitation was done using highly selective and sensitive liquid chromatography-tandem mass spectrometry, with a limit of quantitation at 2.2 lesions per 10
Identifiants
pubmed: 31055913
doi: 10.1021/acs.analchem.9b01351
pmc: PMC6718090
mid: NIHMS1047576
doi:
Substances chimiques
DNA
9007-49-2
DNA-(Apurinic or Apyrimidinic Site) Lyase
EC 4.2.99.18
Types de publication
Journal Article
Research Support, N.I.H., Extramural
Langues
eng
Sous-ensembles de citation
IM
Pagination
7403-7410Subventions
Organisme : NCI NIH HHS
ID : P01 CA160032
Pays : United States
Organisme : NCI NIH HHS
ID : P30 CA077598
Pays : United States
Organisme : NCATS NIH HHS
ID : UL1 TR000114
Pays : United States
Organisme : NCI NIH HHS
ID : P30 CA068485
Pays : United States
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