The Single Cell Gel Assay: A Sensitive Technique for Evaluating Intercellular Differences in DNA Damage and Repair

Abstract

While providing information at the level of the individual cell, cytogenetic techniques for evaluating DNA damage or repair are, by their very nature, largely limited to proliferating cell populations. Furthermore, these techniques require the processing of DNA damage into microscopically visible lesions. Biochemical techniques, such as alkaline elution and nucleoid sedimentation, circumvent these difficulties in that DNA damage can be evaluated directly in almost any cell population. However, the resulting data do not provide any information about the distribution of damage or repair among individual cells. Since the effects of genotoxic agents are often tissue and cell-type specific, techniques which can directly detect DNA damage in individual cells are needed. Rydberg and Johanson (1978) were the first to directly quantitate DNA damage in individual cells by lysing cells embedded in agarose on slides under mild alkali conditions to allow the partial unwinding of DNA. After neutralization, the cells are stained with acridine orange and the extent of DNA damage quantitated by measuring the ratio of green (indicating double-stranded DNA) to red (indicating single-stranded DNA) fluorescence using a photometer.