Abstract
Apurinic/apyrimidinic (AP) sites are produced as the intermediates in the course of the base excision repair (BER) via several specific DNA glycosylases. We have recently developed a method for detection and quantitation of AP sites in DNA via Aldehyde Reactive Probe (ARP) which specifically reacts with aldehyde group of AP sites. In this study, we have directly analyzed intracellular AP sites using Fluorescein Aldehyde Reactive Probe-1 (FARP-1, DOJINDO Laboratories). After incubating HeLa RC355 cells with 0, 2.5 and 5.0 mM methylmethanesulfonate (MMS) for 1 hour to produce AP sites, cells were harvested, treated in Triton X-100 and RNase A/PBS(-) solution, and then stained with FARP-1 for 2 hours. They were analyzed by a flowcytometer. The signal intensities were constant at the concentrations more than 0.1 mM. A linear relationship between MMS concentrations and the signal intensities was observed. By establishing this new method, it will be possible to investigate in situ repair process of DNA damages in the cell. [J Radiat Res 44:437 (2003)]
