Joint Research Activities
This work package will be led by CNRS because of their experience in the biomedical applications of ion beam techniques. They also lead the key task, which is directed towards single ion irradiation of living cells in medium. Another keytask capitalises on the experience of CEA in the development of novel in situ measurement methodologies using ion beams. All of the developments in this work package will occur at infrastructures offering transnational access. Consequently as these techniques are developed they will be made available to the wider European community and maintain and enhance the excellence and world class reputation of the TNA providers.
Key task: Targeted Irradiation
The advent of targeted irradiation techniques allowing the delivery of precisely counted numbers of ions at pre-determined positions in the sample, on the micron scale, has opened up a large number of applications not only in materials and semiconductor sciences but also in radiation biology. The interest in radiation biology comes from the possibility of irradiating individual living cells in medium with an exact number of ions (down to the ultimate dose of one ion per cell) at specific locations within the cell (for example both nuclear and mitochondrial DNA, the nucleus and cytoplasm themselves, and other cellular organelles and the cell membrane) and then to studying the biological response to DNA or protein damage (signalling pathways, mechanisms of damage and repair, resistance to radiation, etc.).
Two actions will address the improvement of irradiation techniques:
1) Breaking the micron resolution technical deadlock (CNRS, UBW, SUR, RBI)
The aim is to obtain reliable focussed ion beams at submicron resolution in air or medium which will deliver a pre-determined number of ions to microscopic targets (intra-cellular structures or specific regions in electronic devices) with a targeting accuracy better than a fraction of a micron. To achieve this goal CNRS, UBW, SUR and RBI will undertake joint and complementary technical developments and will disseminate them in order to improve the overall technical level of service offered by the TNA facilities.
This will be achieved by:
- Development of new high demagnification multistage optics (UBW, RBI) and commissioning of a original vertical beam line at SUR
- Design of new very thin transmission detection approaches (via secondary electrons or induced light) for light and heavy ions with specific constraints associated to single event irradiation to enable 100% detection efficiency (CNRS, RBI)
- Development of a pulsed proton beam (< 1 ns pulse duration) with about 100 protons per pulse in 500 nm beam spot in order to observe the effects of low and high LET irradiation (UBW, RBI)
- Development of cell recognition techniques for automatic targeted irradiation of large cell populations
- Development of calibration procedures and quantitative analysis of scintillator patterns to achieve high resolution beam targeting (CNRS, UBW, SUR)
2) Designing new approaches to improve the quality of irradiation (CNRS, UBW, SUR)
The reliability of the irradiation procedures
is fundamental in cell radiobiology.
Procedures will be set up to check the efficiency
of single event control (for dose
calculation) and the targeting precision in
Live cell experiments will be developed to detect DNA double strand breaks in-situ and the development of the damage and repair mechanisms subsequent to irradiations (CNRS, UBW, SUR).
Key task : In situ Instrumentation
1) Implementing advanced imaging techniques on beamlines:
In Radiation Biology, the advent of microbeam
irradiation techniques has introduced
a huge potential for monitoring
cellular and molecular events in response
to DNA/chromatin damage.
SPIRIT will increase the implementation of these techniques to other TNA providers in order to improve the access to in situ experiments on living cells via the following actions:
- Comparing advanced imaging techniques to investigate in situ molecular signalling pathways and protein dynamics in the ion tracks, i.e. Fluorescence life-time, wide field versus laser scanning, Fluorescence Recovery After Photo-bleaching (FRAP) techniques (CNRS, UBW, SUR)
- Development of cell lines which incorporate GFP-tagged proteins in order to follow repair kinetics and dynamics of damaged DNA structures. This development is an essential prerequisite (UBW, CNRS)
- Translational research – studying clinically important factors such as the effects of hypoxia and hemotherapeutic
agents (e.g. temozolomide) on the radiation sensitivity of cell populations (SUR).
2) Installing unique on line characterisation instruments
In this key task in situ experiments will be enabled via the following actions:
- IBA with ultra high-resolution will be made available for the analysis of deposited nanoscale thin films (HZDR)
- A real-time in situ characterization chamber for RBS measurement at variable sample temperature will be installed (KUL)
- Luminescence of excitons and radiation-induced defects are also frequently quenched at low temperature and
requires cryogenic devices. Time resolved luminescence on the ns timescale is planned. This instrument will be installed at CEA (IRRSUD) and will be complementary to existing on-line infrared spectroscopy instruments and
the future on-line x-ray powder scattering instrument.