The Induction and Repair of Double-Strand DNA Breaks in Mammalian Cells as Detected by Neutral Elution
One of the important types of damage induced by ionizing radiation in intracellular DNA is the double-strand break (dsb). This damage, involving disruption of both strands of the DNA-helix at the same or neighbouring sites, is produced at a much lower frequency than other lesions (single-strand breaks, nucleotide damage). One would expect, however, that double-strand breaks would be much more deleterious for the cell than lesions of the latter type as these probably are repaired more easily.
The neutral filter elution technique has proven to be a sensitive means of detecting DNA dsb after exposure to low doses of ionizing radiation (Bradley and Kohn, 1979). However, the results obtained with it reported in literature are in some cases controversial as well as in disagreement with results of other methods. For example, according to data of Radford (1985) and Radford and Hodgson (1985), the induction of dsb by 250 kV X-rays does not show a simple linear dose-effect relationship, whereas others (Ross and Bradley, 1981, Van der Schans et al. 1982a, and Woods et al. 1982) found data consistent with a linear relationship. Also Blöcher (1982) observed a linear relationship in his sedimentation studies.
Another discrepancy arose when repair of dsb was studied with the elution technique; half-lifes of about 10 min were found, in contrast to the earlier values of 1–2 h obtained in sedimentation experiments. It has been suggested that the fast-repair component seen in filter elution studies might represent repair of DNA single-strand breaks (ssb). This is unlikely since Bradley and Kohn (1979) have found that the different dsb/ssb ratios determined with filter elution for various agents are comparable with those found with other methods. The fact that most of the filter elutions are carried out at rather high pH, could be ruled out as a possible reason for this discrepancy, since we found that elution at neutral pH leads to the same results.
In this paper the reliability of the different methods for the detection of dsb induced by ionizing radiation and other agents will be discussed.
KeywordsStrand Break Chinese Hamster Ovary Cell Elution Procedure Alkaline Elution Fast Repair
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