DNA-repair protein distribution along the tracks of energetic ions.

A simple model of homogenous chromatin distribution in HeLa-cell nuclei suggests that the track of an energetic ion hits 30 nm chromatin fibers with a mean distance of 0.55 mum. To test this assumption, living HeLa-cells were irradiated at the irradiation setup of the ion microprobe SNAKE using the ion beams provided by the Munich 14 MV tandem accelerator. After irradiation, the distribution of 53BP1 protein foci was studied by immunofluorescence. The observed 53BP1 distribution along the tracks of 29 MeV (7)Li ions and 24 MeV (12)C ions differed significantly from the expectations resulting from the simple chromatin model, suggesting that the biological track structure is determined by cell nuclear architecture with higher order organisation of chromatin.

No evidence for a different RBE between pulsed and continuous 20 MeV protons.

To obtain greater insight into the future potential of tumor radiotherapy using proton beams generated from high-intensity lasers, it is important to characterize the ionization quality of the new beams by measuring the relative biological effectiveness (RBE) under conditions where the full dose at one irradiation site will be deposited by a few proton pulses less than 1 ns in duration. HeLa cells attached to a Mylar foil were irradiated with 70 kV X rays to obtain a reference dose-response curve or with 3 Gy of 20 MeV protons at the Munich tandem accelerator (Garching), either using a continuous mode where a cell sample was irradiated within a 100-ms time span or using a pulsed mode where radiation was given in a single proton pulse of about 1 ns. After irradiation cytochalasin B was added; 24 h later cells were fixed and stained with acridine orange and micronuclei were counted. The X-ray dose-response curve for the production of micronuclei in HeLa cells followed a linear-quadratic model. The corresponding RBE values for 20 MeV protons in pulsed and continuous irradiation modes were 1.07 +/- 0.08 and 1.06 +/- 0.10 in the first proton experiment and 1.09 +/- 0.08 and 1.05 +/- 0.11 in the second, respectively. There was no evidence for a difference in the RBE for pulsed and continuous irradiation of HeLa cells with 20 MeV protons.

Three-dimensional hydrogen microscopy in diamond.

A microprobe of protons with an energy of 17 million electron volts is used to quantitatively image three-dimensional hydrogen distributions at a lateral resolution better than 1 micrometer with high sensitivity. Hydrogen images of a <110>-textured undoped polycrystalline diamond film show that most of the hydrogen is located at grain boundaries. The average amount of hydrogen atoms along the grain boundaries is (8.1 +/- 1.5) x 10(14) per square centimeter, corresponding to about a third of a monolayer. The hydrogen content within the grain is below the experimental sensitivity of 1.4 x 10(16) atoms per cubic centimeter (0.08 atomic parts per million). The data prove a low hydrogen content within chemical vapor deposition-grown diamond and the importance of hydrogen at grain boundaries, for example, with respect to electronic properties of polycrystalline diamond.

Microirradiation of cells with energetic heavy ions.

The ion microprobe SNAKE at the Munich 14 MV tandem accelerator achieves beam focussing by a superconducting quadrupole doublet and can make use of a broad range of ions and ion energies, from 20 MeV protons to 200 MeV gold ions. Because of these properties, SNAKE is particularly attractive for biological microbeam experiments. Here we describe the adaptation of SNAKE for microirradiation of cell samples. This includes enlarging of the focal distance in order to adjust the focal plane to the specimen stage of a microscope, construction of a beam exit window in a flexible nozzle and of a suitable cell containment, as well as development of procedures for on-line focussing of the beam, preparation of single ions and scanning by electrostatic deflection of the beam. When irradiating with single 100 MeV (16)O ions, the adapted set-up permits an irradiation accuracy of 0.91 microm (full width at half maximum) in the x-direction and 1.60 microm in the y-direction, as demonstrated by retrospective track etching of polycarbonate foils. Accumulation of the repair protein Rad51, as detected by immunofluorescence, was used as a biological track detector after irradiation of HeLa cells with geometric patterns of counted ions. Observed patterns of fluorescence foci agreed reasonably well with irradiation patterns, indicating successful adaptation of SNAKE. In spite of single ion irradiation, we frequently observed split fluorescence foci which might be explained by small-scale chromatin movements.