U.S. flag An official website of the United States government.
Official websites use .gov

A .gov website belongs to an official government organization in the United States.

Secure .gov websites use HTTPS

A lock ( ) or https:// means you've safely connected to the .gov website. Share sensitive information only on official, secure websites.

i

Importance of Complete DNA Digestion in Minimizing Variability of 8-Oxo-dG Analyses



Details

  • Personal Author:
  • Description:
    Estimates of 8-oxo-2'-deoxyguanosine (8-oxo-dG) in DNA vary at least one order of magnitude using different quantitative methods or even the same method. Our hypothesis is that an incomplete DNA hydrolysis to nucleosides by the conventional nuclease P1 (NP1) and alkaline phosphatase (AP) digestion system plays an important role in contributing to the variability of measurements using HPLC coupled with UV and electrochemical (EC) detection. We show here that factors, such as the amount of DNA, choice of enzymes, their activities, and incubation time, can affect DNA digestion and, thus, cause variability in 8-oxo-dG levels. The addition of DNase I and phosphodiesterases I and II to the NP1 + AP system improves the DNA digestion by completely releasing normal nucleosides and 8-oxo-dG, thereby reducing the interday variations of 8-oxo-dG levels. Diethylenetriamine pentaacetic acid (DTPA), an iron chelator, prevented background increases of 8-oxo-dG during DNA digestion, as well as during the waiting period in the autosampler when a batch of DNA samples is analyzed by HPLC. After optimization of the DNA digestion conditions, the interday variability of 8-oxo-dG measurements using commercially available salmon testes DNA (ST DNA) were 26% over a period of 2 years. Under these optimal conditions, our laboratory variability may contribute as little as 13% to the overall variability as shown by assessment of oxidative DNA damage in a population of smokers. Based on our results, we believe that the modified DNA digestion conditions will provide much more accurate 8-oxo-dG determinations and, thus, more reliable estimates of cancer risk. [Description provided by NIOSH]
  • Subjects:
  • Keywords:
  • ISSN:
    0891-5849
  • Document Type:
    Text
  • Funding:
    Grant
  • Genre:
    Journal Article
  • Place as Subject:
  • CIO:
    National Institute for Occupational Safety and Health (NIOSH)
  • Topic:
    Workplace Safety & Health
  • Location:
    North America
  • Volume:
    31
  • Issue:
    11
  • NIOSHTIC Number:
    nn:20057884
  • Citation:
    Free Radic Biol Med 2001 Dec; 31(11):1341-1351
  • Contact Point Address:
    Dr. Krystyna Frenkel, New York University School of Medicine, Department of Environmental Medicine, PHL Room 802, 550 First Avenue, New York, NY 10016, USA
  • Email:
    krystyna.frenkel@nyu.edu
  • Federal Fiscal Year:
    2002
  • Performing Organization:
    New York University Medical Center, New York, New York
  • Peer Reviewed:
    True
  • Start Date:
    19990930
  • Source Full Name:
    Free Radical Biology and Medicine
  • End Date:
    20030929
  • Collection(s):
  • Main Document Checksum:
    urn:sha-512:dec520a30b3e9ebb58bb1d1bcd9895323e12c8e8568061fb27576b6bdf7ba8ee7f1491648f8407adb2acdeb744c46ac4baad6496cae39596a7c6ee41c52deed6
  • Download URL:
  • File Type:
    Filetype[PDF - 220.57 KB ]
PREVIOUS
ON THIS PAGE
Details
CDC STACKS serves as an archival repository of CDC-published products including scientific findings, journal articles, guidelines, recommendations, or other public health information authored or co-authored by CDC or funded partners.

As a repository, CDC STACKS retains documents in their original published format to ensure public access to scientific information.
BACK TO TOP