Photobleaching setup for the biological end-station of the darmstadt heavy-ion microprobe

We report the upgrade of the epifluorescence microscope of the GSI heavy-ion microprobe with a galvo-scanned, 488 nm laser diode. The laser is focussed into the object plane by the water-immersion objective resulting in a focal spot size of about 1 μm. To increase temporal and spatial resolution a water-immersion objective with a high numerical aperture is integrated into the custom-build microscope. The upgraded system can now be used to bleach GFP-tagged proteins recruited to DNA damage induced by targeted single-ion irradiation. The system is demonstrated on NIH 3T3 cells with Ku80-GFP ion-targeted in heterochromatic and euchromatic DNA. Fluorescence recovery after photobleaching (FRAP) is shown to be significantly slower in heterochromatin.