Application of the micro-PIXE technique in biomedical research

Preparation of samples for micro-PIXE analysis on biological tissue or individual cells is a demanding process, which dominantly influences the quality of the results. Shock-freezing, cryotome slicing and freeze-drying of the tissues are most frequently used steps for the tissue preparation. However, the removal of water induces morphological alterations at the subcellular level. A need for preserving the sub-cellular morphology motivated the efforts to keep the tissue in a frozen hydrated state during the analysis. First micro-PIXE measurements on frozen hydrated tissues were reported in 2007 by the iThemba group [1]. Several micro-XRF facilities at synchrotrons started to offer such sample handling for external users. We report on instrumental and methodological development of micro-PIXE analysis on frozen hydrated tissue at JSI.

After the incorporation of a multicusp ion source at JSI tandem accelerator, high-energy proton beam brightness is increased for one order of magnitude in comparison with earlier used duoplasmatron ion source [1]. This results in the availability of sub-micrometer beam sizes for micro-PIXE (600 nm) and correspondingly the sub-cellular resolution in the tissue mapping.

Several recent applications of micro-PIXE at JSI are addressing plant physiology [2], nanomedicine [3], nanotoxicology [4], food research [5] and environmental pollution [6].

Figure 1. Example: wheat grains exposed to moisture for 2 hours, shock-freezed, freeze-dried, munted on Al holders within two pioloform foils, photographed by fluorescence microscope (Collaboration with K. Vogel Mikuš, Biotechnical faculty, University of Ljubljana).

Figure 2. Parallel acquisition of five spectra in a list mode (OmDaq)

Figure 3. PIXE maps accumulated, OmDAQ, Proton energy 2.5 MeV, overnight

Figure 4. Listmode files processed by GEOPIXE program (C. Ryan)

Figure 5. Zoom on aleurone

Figure 6. Listmode files processed by GEOPIXE program

Figure 7. Last developments: Micro-PIXE on frozen hydrated biological tissue

References:

  1. G. Tylko et al, Nucl. Instr. Meth. B 260 (2007) 141.
  2. P. Vavpetič et al, Nucl. Instr. Meth. B 306 (2013) 140.
  3. P. Pelicon et al, Nucl. Instr. Meth. B, 2014, DOI:10.1016/j.nimb.2014.02.067.
  4. I. Lefevre et al, Plant, Cell & Environm., 37 (2014), 1299.
  5. S. Tomić et al, accepted for publ. in PLOS ONE.
  6. S. Novak et al, Environ. Sci. & Tech. 47 (2013), 5400.
  7. P. Pongrac et al, J. R. Soc. Interface 84 (2013), 1742.
  8. L. Lyubenova et al, J. Haz. Mat. 248/249 (2013), 371.