Microbeam design in radiobiological research

Abstract

Recent work using low-doses of ionising radiations, both in vitro and in ViVO, has suggested that the responses of biological systems in the region of less than 1 Gray may not be predicted by simple extrapolation from the responses at higher doses. Additional experiments, using high-LET radiations at doses of much less than one alpha particle traversal per cell nucleus, have shown responses in a greater number of cells than have received a radiation dose. These findings, and increased concern over the effects of the exposure of the general population to low-levels of background radiation, for example due to radon daughters in the lungs, have stimulated the investigation of the response of mammalian cells to ionising radiations in the extreme low-dose region. In all broad field exposures to particulate radiations at low-dose levels an inherent dose uncertainty exists due to random counting statistics. This dose variation produces a range of values for the measured biological effect within the irradiated population, therefore making the elucidation of the dose-effect relationship extremely difficult. The use of the microbeam irradiation technique will allow the delivery of a controlled number of particles to specific targets within an individual cell with a high degree of accuracy. This approach will considerably reduce the level of variation of biological effect within the irradiated cell population and will allow low-dose responses of cellular systems to be determined. In addition, the proposed high spatial resolution of the microbeam developed will allow the investigation of the distribution of radiation sensitivity within the cell, to provide a better understanding of the mechanisms of radiation action. The target parameters for the microbeam at the Gray Laboratory are a spatial resolution of less than 1 urn and a detection efficiency of better than 99 %. The work of this thesis was to develop a method of collimation, in order to produce a microbeam of 3.5 MeV protons, and to develop a detector to be used in conjunction with the collimation system. In order to determine the optimum design of collimator necessary to produce a proton microbeam, a computer simulation based upon a Monte-Carlo simulation code, written by Dr S J Watts, was developed. This programme was then used to determine the optimum collimator length and the effects of misalignment and divergence of the incident proton beam upon the quality of the collimated beam produced. Designs for silicon collimators were produced, based upon the results of these simulations, and collimators were subsequently produced for us using techniques of micro-manufacturing developed in the semiconductor industry. Other collimator designs were also produced both in-house and commercially, using a range of materials. These collimators were tested to determine both the energy and spatial resolutions of the transmitted proton beam produced. The best results were obtained using 1.6 mm lengths of 1.5 µm diameter bore fused silica tubing. This system produced a collimated beam having a spatial resolution with 90 % of the transmitted beam lying within a diameter of 2.3 ± 0.9 µm and with an energy spectrum having 75 % of the transmitted protons within a Gaussian fit to the full-energy peak. Detection of the transmitted protons was achieved by the use of a scintillation transmission detector mounted over the exit aperture of the collimator. An approximately 10 urn thick ZnS(Ag) crystal was mounted between two 30 urn diameter optical fibres and the light emitted from the crystal transmitted along the fibres to two photomultiplier tubes. The signals from the tubes were analyzed, using coincidence counting techniques, by means of electronics designed by Dr B Vojnovic. The lowest counting inefficiencies obtained using this approach were a false positive count level of 0.8 ± 0.1 % and an uncounted proton level of 0.9 ± 0.3 %. The elements of collimation and detection were then combined in a rugged microbeam assembly, using a fused silica collimator having a bore diameter of 5 urn and a scintillator crystal having a thickness of - 15 µm. The microbeam produced by this initial assembly had a spatial resolution with 90 % of the transmitted protons lying within a diameter of 5.8 ± 1.6 µm, and counting inefficiencies of 0.27 ± 0.22 % and 1.7 ± 0.4 % for the levels of false positive and missed counts respectively. The detector system in this assembly achieves the design parameter of 99 % efficiency, however, the spatial resolution of the beam is not at the desired I urn level. The diameter of the microbeam beam produced is less than the nuclear diameter of many cell lines and so the beam may be used to good effect in the low-dose irradiation of single cells. In order to investigate the variation in sensitivity within a cell the spatial resolution of the beam would require improvement. Proposed methods by which this may be achieved are described.