Single Ion Hit Facility Research Articles

Live cell calcium imaging at the single ion hit facility of GSI

To study the fast intracellular calcium response after ion irradiation in living mammalian cells, a live cell calcium imaging set-up was constructed at the targeted cell irradiation facility at GSI. This work introduces the live cell calcium imaging system, shows its performance, an example of the ratio-metric calcium measurement and its application to on-line study calcium response to targeted ion irradiation in human cells.

Applications of SU-8 in the development of a Single Ion Hit Facility

The Lund Nuclear Microprobe (LNM) has been adapted to be used as a Single Ion Hit Facility (SIHF) for proton cell irradiation experiments at low dose. In order to test the capabilities of the system, Ni dot arrays and artificial cells have been fabricated with the photopolymer SU-8 and common lithographic techniques. The primary purpose of the Ni dot arrays was to determine the targeting accuracy of the beam in vacuum and in air. Additionally, this sample was employed to evaluate the system performance during cell target irradiation experiments. The Ni dot arrays were also used for beam characterization. The artificial cells were originally fabricated to test the software for cell recognition and localization, developed and implemented at the LNM. However, this sample became very functional to make small adjustments at the irradiation chamber for cell irradiation experiments. A description of the samples, fabrication procedure and applications are presented in this paper.

The Absence of an Early Calcium Response to Heavy-Ion Radiation in Mammalian Cells

Intracellular calcium is an important second messenger that regulates many cell functions. Recent studies have shown that calcium ions can also regulate the cellular responses to ionizing radiation. However, previous data are restricted to cells treated with low-LET radiations (X rays, gamma rays and beta particles). In this work, we investigated the calcium levels in cells exposed to heavy-ion radiation of high LET. The experiments were performed at the single ion hit facility of the GSI heavy-ion microprobe. Using a built-in online calcium imaging system, the intracellular calcium concentrations were examined in HeLa cells and human foreskin fibroblast AG1522-D cells before and after irradiation with 4.8 MeV/nucleon carbon or argon ions. Although the experiment was sensitive enough to detect the calcium response to other known stimuli, no response to heavy-ion radiation was found in these two cell types. We also found that heavy-ion radiation has no impact on calcium oscillation induced by hypoxia stress in fibroblast cells.

Double strand break formation as a response to X-ray and targeted proton-irradiation

The use of targeted ion beams has been an important development for understanding the response of biological systems to radiation exposure. Most investigations in this field have been performed with helium ion microbeams; however, irradiations using a beam of protons, covering different LET values, can deliver complementary results and additional information on radiation effect related to radiation quality and low dose exposure. Recently, several experiments have been carried out at the new single ion hit facility in the Institute of Nuclear Physics in Cracow using a focused proton microbeam. In parallel, similar cross-validation experiments have been performed using the well-established particle microbeam situated at the Gray Cancer Institute with the same cell line and proton energy. Pilot studies have compared the yields of double strand breaks (DSBs) in cells uniformly irradiated with 240 kV X-ray with those irradiated with microbeam delivered protons, and in some cases helium ions. The number of DSBs increased with the X-ray dose. In the samples irradiated with individually counted protons, fewer foci were found in comparison to the number of protons delivered per cell. Studies also compared the distribution of foci per cell between the different radiation qualities.

The new Cell Irradiation Facility at the Lund Nuclear Probe

The CELLION project is directed towards the studies on cellular response to targeted single ions. This paper gives an account of the modifications made at the Lund sub-micron beam line in order to create a Single Ion Hit Facility for biological applications within the CELLION project. The most relevant new feature is the specially developed software used to locate the cells. A program for cell recognition and localization that gives the coordinates of the centre of the cells has been designed. The recognition is made online. A picture taken by a microscope is used as input parameter for the recognition program. Using V79 Chinese hamster cells, the recognition procedure can be done in less than 0.5 s for a picture size of 800 × 600 pixels with approximately 96% efficiency. This paper also reports target accuracy test results and the first non-targeted irradiation procedure performed at the Lund sub-micron beam line.

Development of the IFJ single ion hit facility for cell irradiation

In recent years, a single ion hit facility has been constructed at the IFJ ion microprobe. The setup is used for the precise irradiations of living cells by a controlled number of ions. Investigations of such type have two very important requirements: (1) cells must be examined in their natural state and environment (i.e. without previously being killed, preferentially neither fixed nor stained) and (2) the possibility of automatic irradiation of large number of cells (including computer recognition of cells positions) must be provided. This work presents some of the crucial features of the off-line and on-line optical systems, including self-developed software responsible for automatic cell recognition.

Development of a single ion hit facility at the Pierre Süe Laboratory: a collimated microbeam to study radiological effects on targeted living cells

A single ion hit facility is being developed at the Pierre Süe Laboratory (LPS) since 2004. This set-up will be dedicated to the study of ionising radiation effects on living cells, which will complete current research conducted on uranium chemical toxicity on renal and osteoblastic cells. The study of the response to an exposure to alpha particles will allow us to distinguish radiological and chemical toxicities of uranium, with a special emphasis on the bystander effect at low doses. Designed and installed on the LPS Nuclear microprobe, up to now dedicated to ion beam microanalysis, this set-up will enable us to deliver an exact number of light ions accelerated by a 3.75 MV electrostatic accelerator. An 'in air' vertical beam permits the irradiation of cells in conditions compatible with cell culture techniques. Furthermore, cellular monolayer will be kept in controlled conditions of temperature and atmosphere in order to diminish stress. The beam is collimated with a fused silica capillary tubing to target pre-selected cells. Motorisation of the collimator with piezo-electric actuators should enable fast irradiation without moving the sample, thus avoiding mechanical stress. An automated epifluorescence microscope, mounted on an antivibration table, allows pre- and post-irradiation cell observation. An ultra thin silicon surface barrier detector has been developed and tested to be able to shoot a cell with a single alpha particle.

A fast online hit verification method for the single ion hit system at GSI

For a single ion hit facility built to irradiate specific targets inside biological cells, it is necessary to prove that the ions hit the selected targets reliably because the ion hits usually cannot be seen. That ability is traditionally tested either indirectly by aiming at pre-etched tracks in a nuclear track detector or directly by making the ion tracks inside cells visible using a stain coupled to special proteins produced in response to ion hits. However, both methods are time consuming and hits can be verified only after the experiment. This means that targeting errors in the experiment cannot be corrected during the experiment. Therefore, we have developed a fast online hit verification method that measures the targeting accuracy electronically with a spatial resolution of +/-1 microm before cell irradiation takes place.

Evaluation of a pre-cell hit detector for the future single ion hit facility in Lund

Currently, a single ion hit facility, for irradiation of single cells with single, light MeV ions is under development at the Lund Nuclear Microprobe. In this paper, a novel approach to the ion detection issue is presented. A silicon detector, a type utilized at other facilities for post-cell ion detection, has been investigated as a possible option for pre-cell hit detection. If proven successful, this detector could possibly also be used simultaneously as vacuum window.The first experiments carried out on the 9μm thick silicon detector, with an area of 4mm2, have been aimed at investigations of signal-to-noise ratio and efficiency. The results thus far reveal a low noise level and a noise distribution, which is well separated from the signal peak. However, the efficiency remains a problem, since at present it is far from the required 100%.

Irradiating single cells using Cracow microprobe facility

The Cracow single ion hit facility (SIH) is using the Van de Graaff-based microprobe with current spatial resolution of about 10μm in vacuum. Living cells are irradiated with the exact number of 2MeV protons. This work shows the present setup and test measurements optimizing the detection of single protons, beam deflection after registering single proton and the accuracy of cell targeting as well as the description of performance of the whole setup. The article presents also ways of cells visualisation and positioning during a single measurement and reports on first experiments with living cells carried out at the new facility.

Automatic system for single ion/single cell irradiation based on Cracow microprobe

Recently, the Cracow ion microprobe has found its new application as a single ion hit facility (SIHF), allowing precise irradiations of living cells by a controlled number of ions. The instrument enables a broad field of research, such as survival studies, adaptive response investigations, bystander effect, inverse dose-rate effect, low-dose hypersensitivity, etc. This work presents principles of construction and operation of the SIHF based on the Cracow microprobe. We discuss some crucial features of optical, positioning, and blanking systems, including self-developed software responsible for semiautomatic cell recognition, for precise positioning of cells, and for controlling the irradiation process. We also show some tests carried out to determine the efficiency of the whole system and of its segments. In addition, we present results of the first irradiation measurements performed with living cells.

The GSI Heavy Ion Microbeam: A Tool for the Investigation of Cellular Response to High LET Radiations

Since the mid 1990’s, an increasing number of charged particle microbeams have been designed to deliver a preset number of ions to individual living cells with the micron resolution. These tools provide a powerful technique to investigate the cellular response to low doses of radiations. During the last years, the single ion hit facility in operation on the GSI microbeam since 1987 has been upgraded for the irradiation of individual living cells in vitro. This setup presents two main peculiarities compared to the microbeams used up to now for cell irradiation. First, the beam’s micrometric size is obtained by magnetic focusing and not by a simple collimation. This allows obtaining a smaller beam spot, a better defined linear energy transfer, and a high irradiation throughput. Then, the GSI microbeam is able to focus ions from carbon to uranium with energies between 1.4 MeV/u to 11.4 MeV/u. The range of accessible linear energy transfer is thus considerably extended compared to light ions microbeam in operation today. The design of the GSI microbeam is described, including the beam control, the online cell localisation, the cell dish designed specifically for microbeam irradiation, and the cell irradiation procedures. Experimental tests performed to check the global aiming accuracy as well as the first cellular irradiations are presented.

Preparatory experiments for a single ion hit facility at GSI

This article describes results of a continuing effort in development of a “second generation” biological single ion hit facility (SIHF) at GSI, Darmstadt, Germany. It describes the rationale, developmental details and current status of the SIHF at GSI.

Design of a single ion hit facility

The aim of this project is to develop a new system for single ion irradiation of cells for genetic and cell biology studies. This charged particle focused microbeam system will provide a fully computer controlled irradiation facility with submicron (subcellular) resolution and will open many new avenues into studies of radiobiological mechanisms. In the first stage of this project a system for single ion detection has been developed. A thin diamond window has been tested as vacuum/atmosphere window and as a possible source of secondary electrons and/or photons for single ion detection. A detection efficiency of 97% has been achieved.