Radiation-induced DNA single-strand breaks in freshly isolated human leukocytes

J. M. Buatti, L. R. Rivero, T. J. Jorgensen

Research output: Contribution to journalArticle

Abstract

Single-strand breaks are a major form of DNA damage caused by ionizing radiation, and measurement of strand breaks has long been used as an index of overall cellular DNA damage. Most assays for DNA single-strand breaks in cells rely on measuring fractionated DNA samples following alkali denaturation. Quantification is usually achieved by prelabeling cells with radioactive DNA precursors; however, this is not possible in the situation of nondividing cells or freshly isolated tissue. It has previously been demonstrated that the alkali unwinding assay of DNA strand breaks can be quantified by blotting the recovered DNA on nylon membranes and hybridizing with radiolabeled sequence-specific probes. We report here improvements to the technique, which include hot alkali denaturation of DNA samples prior to blotting and the use of carrier DNA that is non-complementary to the radiolabeled probe. Our method allows both single- and double-stranded DNA to be quantified with the same efficiency, thereby improving the sensitivity and reproducibility of the assay, and allows calibration for determination of absolute levels of DNA strand breaks in cells. We also used this method to assay radiation-induced DNA strand breaks in freshly isolated human leukocytes and found them to have a strand break induction rate of 1815 strand breaks/cell/Gy.

Original languageEnglish (US)
Pages (from-to)200-206
Number of pages7
JournalRadiation Research
Volume132
Issue number2
DOIs
StatePublished - 1992
Externally publishedYes

Fingerprint

Single-Stranded DNA Breaks
leukocytes
strands
Leukocytes
deoxyribonucleic acid
DNA Breaks
Radiation
Alkalies
DNA
radiation
DNA Damage
alkalis
Nucleic Acid Denaturation
Single-Stranded DNA
Nylons
assays
alkalies
cells
Ionizing Radiation
denaturation

ASJC Scopus subject areas

  • Agricultural and Biological Sciences (miscellaneous)
  • Radiology Nuclear Medicine and imaging
  • Biophysics
  • Radiation

Cite this

Radiation-induced DNA single-strand breaks in freshly isolated human leukocytes. / Buatti, J. M.; Rivero, L. R.; Jorgensen, T. J.

In: Radiation Research, Vol. 132, No. 2, 1992, p. 200-206.

Research output: Contribution to journalArticle

Buatti, J. M. ; Rivero, L. R. ; Jorgensen, T. J. / Radiation-induced DNA single-strand breaks in freshly isolated human leukocytes. In: Radiation Research. 1992 ; Vol. 132, No. 2. pp. 200-206.
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