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Philosophical Transactions of the Royal Society B: Biological Sciences logoLink to Philosophical Transactions of the Royal Society B: Biological Sciences
. 1998 Jun 29;353(1370):895–902. doi: 10.1098/rstb.1998.0253

Analysis of DNA damage and repair accompanying differentiation in the intestinal crypt.

D J Winton 1, R A Brooks 1
PMCID: PMC1692273  PMID: 9684286

Abstract

The ability to process damaged DNA may vary between cells depending on their differentiated status. However, there is little in vivo data available and it is not intuitively obvious how the activity of specific repair pathways may vary between different subpopulations (e.g. stem cells and proliferative, committed and differentiated cells) of a particular tissue. To obtain such information for the intestinal epithelium, we have developed an assay that detects differences in the way different regions of the crypt (stem, proliferative and maturation zones) respond to DNA damage. The assay is a variant of the 'comet' assay, which detects DNA strand breaks by measuring the proportion of DNA migrating from individual cells, or in this case intact isolated crypts, in an electrophoretic field. The method is quantitative, with the amount of migrating DNA being proportional to the number of strand breaks. Isolated crypts are repair competent and spatial differences are apparent with some agents. The assay has the potential to characterize the repair properties of cells at different stages of differentiation within the crypt, determine the characteristics that might predispose them to damage and may help in understanding the route of stem cell mutation.

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Selected References

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