DNA microfiltration assay: a simple technique for detecting DNA damage in mammalian cells

A simple method for detection of DNA single-strand breaks (DNA-SSB) in cultured cells is described, based on filtration of alkaline-lysed cells through microfilters. After exposure to potentially DNA damaging agents, the cells are transferred to 0.8 micron cellulose acetate filters mounted in microfilter devices where they are washed, lysed and centrifuged to separate undamaged DNA from damaged DNA. When human bronchiolar cells (14Br) were exposed to different DNA damaging agents, hydrogen peroxide, N-methyl-N-nitro-N-nitrosoguanidine or 4-nitroquinoline-1-oxide, there was good correlation between the extent of DNA damage assessed by this filtration technique and by DNA precipitation assay. DNA-SSB were also detected by the filtration technique after exposure of bronchiolar cells to phorbol ester-stimulated human neutrophils. The filtration assay is easy to perform, the sample handling capacity is very high, and no expensive or complicated laboratory equipment is required. It may therefore be an alternative, or a complement, to other methods for detection of DNA-SSB.