Measurement of DNA damage in mammalian cells using flow cytometry

A. E. Milner, Andrew T M Vaughan, I. P. Clark

Research output: Contribution to journalArticle

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Abstract

A technique for the detection of DNA damage induced by radiation insult has been developed. Cells were lysed with a buffer containing 2 M sodium chloride to release the DNA in a supercoiled from, the nucleoid. These were stained with the DNA intercalating dye, ethidium bromide, and exposed to laser light within a flow cytometer. Scattered and fluorescent light was analyzed form the laser/nucleoid interaction following irradiation of viable cells with γ rays. The addition of ethidium bromide to prepared nucleoids caused a reduction in scattered light due to condensation of the nucleoid. Irradiation of cells prior to nucleoid production and ethidium bromide treatment restricted this condensation and produced a dose-dependent increase in laser scatter. Nucleoids derived from human lymphocytes showed enhanced light scatter from 5 Gy, compared to Chinese hamster ovary (CHO) fibroblasts where doses above 10 Gy were required. Up to 30 Gy CHO nucleoids showed a dose-dependent reduction in the ethidium bromide fluorescence. This technique allows detection of altered light scattering and fluorescent behavior of nucleoids after cellular irradiation; these may be related to structural changes within the nucleus induced by the radiation. The use of flow cytometry compared to other methods allows a rapid analysis of nuclear damage within individual cells.

Original languageEnglish (US)
Pages (from-to)108-117
Number of pages10
JournalRadiation Research
Volume110
Issue number1
StatePublished - 1987
Externally publishedYes

Fingerprint

cytometry
DNA damage
Ethidium
DNA Damage
flow cytometry
bromides
Flow Cytometry
deoxyribonucleic acid
damage
lasers
Light
ovaries
hamsters
irradiation
Chinese hamsters
Lasers
dosage
Cricetulus
condensation
cells

ASJC Scopus subject areas

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

Cite this

Measurement of DNA damage in mammalian cells using flow cytometry. / Milner, A. E.; Vaughan, Andrew T M; Clark, I. P.

In: Radiation Research, Vol. 110, No. 1, 1987, p. 108-117.

Research output: Contribution to journalArticle

Milner, A. E. ; Vaughan, Andrew T M ; Clark, I. P. / Measurement of DNA damage in mammalian cells using flow cytometry. In: Radiation Research. 1987 ; Vol. 110, No. 1. pp. 108-117.
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