- Email: [email protected]
The application of stereo scanning transmission ion microscopy (stereo-STIM) imaging in biological specimen
Nuclear Inst. and Methods in Physics Research B xxx (xxxx) xxx–xxx
Contents lists available at ScienceDirect
Nuclear Inst. and Methods in Physics Research B journal homepage: www.elsevier.com/locate/nimb
The application of stereo scanning transmission ion microscopy (stereoSTIM) imaging in biological specimen Ebrahim Gholami Hatam Physics Department, Science Faculty, Malayer University, Malayer, Iran
ARTICLE INFO
ABSTRACT
Keywords: Stereo-STIM Biological sample Disparity
The quantitative two-dimensional areal density can be revealed from the scanning transmission ion microscopy (STIM) image of the sample owing to the relationship between the beam energy loss and the sample areal density. The obtained areal density map can provide fine spatial resolution to identify microstructures. Furthermore, this information can be used to achieve more precise quantification on X-ray absorption if simultaneously performing PIXE analysis. The development of STIM tomography enables to access the inner structure of the sample, especially in studying biological specimens. However, STIM tomography requires a large number of projections while stereo-STIM pair projections provide a quick tool for imaging the shape of the sample and determining the relative position, orientation and shape of the internal and external structures. Stereo-STIM experiments using 2 MeV proton microbeam were carried out on an insect and transmitted energies data were simultaneously collected by two surface barrier detectors mounted on 30° and -30° with respect to the beam direction. The defined disparity value resolved the features of different parts of the body of an insect not seen in individual STIM data.
1. Introduction During the last decade, transmission ion scanning from a simple imaging technique has been developed to a versatile technique of analysis with wide application [1–3] or in combination with other nuclear microprobe techniques such as X-ray analysis induced by incident particle and Rutherford backscattering spectroscopy [4–6]. STIM analysis method can provide density information from different parts of the sample [7] through the simple relationship between beam energy loss and sample areal density. The energy loss is primarily derived from the interaction of ions with electrons and depends on the relative atomic weight, mass density and thickness of the sample according to Bethe-Bloch equation [8]. There are some drawbacks with common set-up/geometry of STIM imaging. In the case of multi-elemental sample having a variety mass density, the yield spectrum from each element has a different forward/ backward scattering kinematic factor then contributing to different energy loss. This will in turn decrease the contrast in the imaging [9]. As a another case, when focused MeV microbeams are scanned over a sample, the energy of the transmitted ions can be measured for each of the defined pixels and it can be used to form 2-D sample energy loss maps [10]. Although these images essentially can be formed from one ion in each pixel. To obtain images with more clarity, a number of counts per pixel are needed. It should be noted that these maps are not
representative of mass density due to the variation of energy loss caused by areal density variations of the sample. There are several ways in which stereoscopic images of ion microprobe can be used to obtain additional information on the sample. Previously, stereo-PIXE was used to quantitatively reconstruct the surface topography of engraved samples [11]. After that, stereoscopic PIXE imaging was displayed as anaglyphs using OMDAQ-3 software [12] to determine the three-dimensional distribution of the micrometeoroids impact in the form of craters from the Hubble Space Telescope [13]. Recently, dual-PIXE tomography system has been used to improve the X-ray detection to reconstruct Germanium in ICF target [14]. Although, stereoscopic STIM (stereo-STIM) imaging has been previously used to provide qualitative three-dimensional density information about certain specimens [15]. In this work, we follow a new approach with the introduction of a “disparity map” in stereo-STIM to generate a correspondence between points of the two images representing the same point of the sample in biological specimen. The use of stereo-STIM could also provide information about spatial distribution and orientation of the small features within a specimen. 2. Stereo-STIM model To study the effect of areal density variation on the result of analysis via stereo-STIM set-up a model of a thin sample with thickness of d is
E-mail address: [email protected]. https://doi.org/10.1016/j.nimb.2018.11.014 Received 11 October 2017; Received in revised form 28 October 2018; Accepted 9 November 2018 0168-583X/ © 2018 Elsevier B.V. All rights reserved.
Please cite this article as: Gholami Hatam, E., Nuclear Inst. and Methods in Physics Research B, https://doi.org/10.1016/j.nimb.2018.11.014
Nuclear Inst. and Methods in Physics Research B xxx (xxxx) xxx–xxx
E. Gholami Hatam
Fig. 1. A schematic layout of stereo-STIM model.
Fig. 2. The energy transmitted spectrum for different Mylar thicknesses. E1, E2 …E5 are the energies transmitted and are plotted by fitting the spectrum to a Gaussian distribution.
Fig. 3. (a) The energy difference versus the energy sum and (b) the disparity value for mean energies indicated at Fig. 2.
considered (Fig. 1). The energy E of a particle after passing through a layer of material with thickness x is given by the integral equation [16] where Ein is the initial energy
The relative areal density variation is embedded in this disparity map. As mentioned, the energy sum at denominator of disparity value is approximately constant for particles scattered at a certain depth while the energy difference in nominator is a fingerprint for inhomogeneity of the sample along the transmitted ions. Thus, true concentration maps can be created to resolve such imaging artefacts that are due to mass density variations.
x
E (x ) = Ein
(dE / dx ') dx '.
(1)
0
The energy sum of the right and left detectors in the case of scattering through 30° can be written as: x
(ER + EL)/2
E
K Ein
(dE / dx )0 dx 0
3. Experimental
3 (d x )/2 '
'
'
'
(dE / dx )1dx .
3.1. Experimental set-up
x
(2)
Using off-axis geometry makes it possible to have enough beam-currents that allow STIM to be used simultaneously with other ion beam analytical techniques such as PIXE and RBS. Stereo-STIM measurements were simultaneously performed with two off-axis STIM set-ups. The two identical particle detectors with resolution of 20 keV, 25 mm2 cross section and 40 mm detector-sample distance, were mounted on 30° and −30° with respect to the beam direction. The protons beam with spatial resolution lower than 10 μm and energy of 2 MeV were scanned over the samples area of 2.4 × 2.4 mm2 and the vacuum chamber pressure was maintained at 1.3 × 10−6 Torr at Van de Graaff accelerator in Tehran [17].
The stopping power, S = (dE/dx), before particles are scattered towards the detector is denoted with “0” and after scattering with “1”. The above equation could also be regarded as the mean of the energy, E , recorded in left and right detectors with respect to the beam direction. It has approximately constant value for particles scattered at depth x, since there is no preferred direction for particles to be scattered at the left or right with the same angle. The scattered protons at the depth of x travel the same distance through the sample, with the result that the relative difference in energy is zero for homogeneous samples; therefore, such events can be selected for further analysis. Accordingly, we have defined the “disparity value” that is implemented for each pixel in the left and right transmitted energy maps with this definition
(E ER ) D= L . (EL + ER )
3.2. Sample preparation For application of the new methodology presented in stereo-STIM an insect was chosen as a biological sample. It was glued to the part of the sample-holder that contains a hole to enhance the imaging and subsequently was air dried.
(3) 2
Nuclear Inst. and Methods in Physics Research B xxx (xxxx) xxx–xxx
E. Gholami Hatam
Fig. 4. (a) and (b) the energy transmitted map for whole body of a flying insect detected in stereo pair of left and right particle detectors. (c) The disparity map calculated from Eq. (3) for every pixel. (d), (e) and (f) are respectively the horizontal profiles of red line indicated in above maps. (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)
3.3. Normalization
i = 1, 2, …, 5. Here, Ē = E3 corresponds to Mylar with areal density of 35,000 (1015 atom/cm2). Fig. 3(b) presents the calculated disparity values of corresponding transmitted energies for known density variations. For discrepancies value close to zero, E ≈ Ē3, we do not have energy difference; hence, the generated disparity map would be null meaning that the sample is homogeneous. The lower (higher) disparity values indicate that the sample is thicker (thinner) than the proposed value of 35,000 (1015 atom/cm2). Interpolation must be used to implement the continuous function given discrete data points in order to determine the mass density variation function.
Regardless the nature of the sample, the value of disparity map in each pixel depends also on the statistic in summed spectra acquired from individual detectors. Even though two particle detectors have had the same geometry and resolution specification the different electronic adjustment may also affect the energy spectrum [18]. In order to consider this effect, a layer of 7 μm Kapton was used to normalize the recorded energy in each pixel of left and right maps. The value of the pixels in the individual energy map was balanced by the total number of particles detected at each detector for Kapton layer.
4.2. Biological sample
4. Result and discussion
As biological samples are mostly suitable to study with STIM analysis a small flying insect was scanned over its whole body. Fig. 4(a) and (b) represent the 2-D average transmitted energy recorded respectively at the left and right detectors. Just considering each of the individual map may ones mislead the interpretation of the result of STIM. Since according to the horizontal profile indicated in the centre of the (Fig. 4(d) and (e)) maps the abdomen part of the body in left detector has more counts with respect to the right spectrometer. At the same time, the head and eyes parts in the right detector have more counts with respect to the left spectrometer. Since the two particle detectors have the same geometry and the maps are normalised, these differences arise from the density variation or different topology along the beam direction that cannot be detected from a single detector spectrometer. Fig. 4(c) represents the disparity map for a whole region of the insect starting from the two point of views on it in the stereo pair. It is indicated from the horizontal profile in the disparity map (Fig. 4(f)) that the features of different parts of the body are resolved which are not seen in individual STIM data. The disparity map matches stereovision and takes the values between −1 (blue color) at the right detector where the left detector is blind and reaches the value of 1 (red color) at the left detector where the other detector is less sensitive.
4.1. Test sample At first, to test the performance of the proposed stereo-STIM model and also to calibrate the areal mass density, different thicknesses of Mylar foils were selected. The beam was scanned over the sample and the energy transmitted from 5 different foils were measured. The accurate mass densities were determined by fitting the corresponding spectra using SIMNRA software [19]. To present the transmitted energy in a two-dimensional map, the average or median energy value of the ions could be calculated in every pixel. Generally, average filtering is better in resolving small structures, even with sub-beam resolution for features of highly varying density [20]. This kind of filtering has been utilised in this study. Fig. 2 shows the spectrum for the transmitted energies corresponding to the 5 different thicknesses of Mylar. Fig. 3(a) illustrates how different mass densities can be resolved from a plot of the energy difference versus the energy sum. It is assumed that if the value of energies measured equals to E = (EL + ER )/2 = E3 and the E3 value is considered to be a reference for defining the change in areal mass densities. Then, the values of Ei − Ē versus Ei + Ē behave linear, where 3
Nuclear Inst. and Methods in Physics Research B xxx (xxxx) xxx–xxx
E. Gholami Hatam
5. Conclusion Application of procedures that can resolve small features within a specimen from a limited number of measurements or reconstruct specimens with non-uniform chemical composition will increase the range of specimens that can be analyzed. Since tomographic investigations are time-consuming and require very small sample dimensions, it could not replace stereo mapping delivering sufficient information in most applications. The stereo-STIM matches the two maps of the transmitted energy in the two detectors by introducing a disparity map. It was concluded that as long as the composition of the sample in known the disparity map is representative of the areal density variation. To validate the proposed model of stereo-STIM different thicknesses of Mylar were examined. In addition, an insect was also analyzed using the proposed stereo-STIM method to resolve different parts of its body. It was also deduced that removing a particular signal from one detector could have the effect of changing the precision of the target areal density as signaled by another detector. In summary, stereo-STIM can be a source of additional information simplifying the interpretation of the data and makes features visible, which cannot be separately distinguished in only two dimensions map.
[4] [5] [6] [7] [8] [9]
[10] [11] [12] [13]
Acknowledgement [14]
This research was supported by Malayer university, contact no. 84/ 9-1-452. I am also greatly indebted to Prof. M. Lamehi-Rachti from Nuclear Science Research School, Tehran, who provided insight and expertise that greatly assisted the research.
[15] [16]
References
[17]
[1] T. Andrea, M. Rothermel, T. Butz, T. Reinert, The improved STIM tomography setup at LIPSION: three-dimensional reconstruction of biological samples, Nucl. Instrum. Meth. Phys. Res. Sect. B 267 (2009) 2098–2102. [2] E.M. Stori, C.T. de Souza, L. Amaral, D. Fink, R.M. Papaléo, J.F. Dias, Use of STIM for morphological studies of microstructured polymer foils, Nucl. Instrum. Meth. Phys. Res. Sect. B 306 (2013) 99–103. [3] P. Aguer, L.C. Alves, P. Barberet, E. Gontier, S. Incerti, C. Michelet-Habchi, Z. Kertész, A.Z. Kiss, P. Moretto, J. Pallon, T. Pinheiro, J.E. Surlève-Bazeille,
[18] [19] [20]
4
Z. Szikszai, A. Veríssimo, M.D. Ynsa, Skin morphology and layer identification using different STIM geometries, Nucl. Instrum. Meth. Phys. Res. Sect. B 231 (2005) 292–299. T. Satoh, T. Sakai, M. Oikawa, Correction of X-ray detection yield in micro-PIXE analysis according to sample thickness variation measured by STIM, Nucl. Instrum. Meth. Phys. Res. Sect. B 249 (2006) 234–237. M.B.H. Breese, G.W. Grime, F. Watt, The use of PIXE and STIM in microelectronics analysis, Nucl. Instrum. Meth. Phys. Res. Sect. B 75 (1993) 341–346. G. Devès, M.P. Isaure, P.L. Lay, J. Bourguignon, R. Ortega, Fully quantitative imaging of chemical elements in Arabidopsis thaliana tissues using STIM, PIXE and RBS, Nucl. Instrum. Meth. Phys. Res. Sect. B 231 (2005) 117–122. I. Bogdanović, S. Fazinić, M. Jakšić, G.W. Grime, V. Valković, Use of STIM in the proton microprobe analysis of single particles, Nucl. Instrum. Meth. Phys. Res. Sect. B 85 (1994) 732–735. S. Tavernier, Interactions of particles in matter, Experimental Techniques in Nuclear and Particle Physics, 2009, pp. 23–53. J. Pallon, V. Auzelyte, M. Elfman, M. Garmer, P. Kristiansson, K. Malmqvist, C. Nilsson, A. Shariff, M. Wegdén, An off-axis STIM procedure for precise mass determination and imaging, Nucl. Instrum. Meth. Phys. Res. Sect. B 219–220 (2004) 988–993. J.C. Overley, R.M.S. Schofield, J.D. Macdonald, H.W. Lefevre, Energy-loss image formation in scanning transmission ion microscopy, Nucl. Instrum. Meth. Phys. Res. Sect. B 30 (1988) 337–341. E.G. Hatam, P. Pelicon, M. Lamehi-Rachti, P. Vavpetič, O. Kakuee, N. Grlj, M. Čekada, V. Fathollahi, Surface topography reconstruction by stereo-PIXE, J. Anal. At. Spectrom. 27 (2012) 834. http://www.microbeams.co.uk/index.html. G.W. Grime, A.T. Kearsley, V. Palitsin, C. Jeynes, J. Colaux, R.P. Webb, Stereoscopic PIXE imaging applied to space debris impact craters from the Hubble Space Telescope, 13th International Conference on Particle Induced X-ray Emission, Gramado (RS), Brazil, (2013). N. Guo, H.Y. Lu, Q. Wang, J. Meng, D.Z. Gao, Y.J. Zhang, X.X. Liang, W. Zhang, J. Li, X.J. Ma, H. Shen, A dual-PIXE tomography setup for reconstruction of Germanium in ICF target, Nucl. Instrum. Methods Phys. B 404 (2017) 162–166. H.W. Lefevre, R.M.S. Schofield, G.S. Bench, G.J.F. Legge, STIM with energy loss contrast: an imaging modality unique to MeV ions, Nucl. Instrum. Meth. Phys. Res. Sect. B 54 (1991) 363–370. C. Wei-Kan, J.W. Mayer, M. Nicolet, Backscattering Spectrometry, ACADEMIC PRESS, INC., New York, 1978. O. Kakuee, V. Fathollahi, M. Lamehi-Rachti, Development of a versatile userfriendly IBA experimental chamber, Nucl. Instrum. Methods Phys. B 371 (2016) 156–160. J.L. Colaux, C. Jeynes, Accurate electronics calibration for particle backscattering spectrometry, Anal. Methods 7 (2015) 3096–3104. M. Mayer, Improved physics in SIMNRA 7, Nucl. Instrum. Meth. Phys. Res. Sect. B 332 (2014) 176–180. G.S. Bench, G.J.F. Legge, High resolution STIM, Nucl. Instrum. Meth. Phys. Res. Sect. B 40–41 (1989) 655–658.
Contents lists available at ScienceDirect
Nuclear Inst. and Methods in Physics Research B journal homepage: www.elsevier.com/locate/nimb
The application of stereo scanning transmission ion microscopy (stereoSTIM) imaging in biological specimen Ebrahim Gholami Hatam Physics Department, Science Faculty, Malayer University, Malayer, Iran
ARTICLE INFO
ABSTRACT
Keywords: Stereo-STIM Biological sample Disparity
The quantitative two-dimensional areal density can be revealed from the scanning transmission ion microscopy (STIM) image of the sample owing to the relationship between the beam energy loss and the sample areal density. The obtained areal density map can provide fine spatial resolution to identify microstructures. Furthermore, this information can be used to achieve more precise quantification on X-ray absorption if simultaneously performing PIXE analysis. The development of STIM tomography enables to access the inner structure of the sample, especially in studying biological specimens. However, STIM tomography requires a large number of projections while stereo-STIM pair projections provide a quick tool for imaging the shape of the sample and determining the relative position, orientation and shape of the internal and external structures. Stereo-STIM experiments using 2 MeV proton microbeam were carried out on an insect and transmitted energies data were simultaneously collected by two surface barrier detectors mounted on 30° and -30° with respect to the beam direction. The defined disparity value resolved the features of different parts of the body of an insect not seen in individual STIM data.
1. Introduction During the last decade, transmission ion scanning from a simple imaging technique has been developed to a versatile technique of analysis with wide application [1–3] or in combination with other nuclear microprobe techniques such as X-ray analysis induced by incident particle and Rutherford backscattering spectroscopy [4–6]. STIM analysis method can provide density information from different parts of the sample [7] through the simple relationship between beam energy loss and sample areal density. The energy loss is primarily derived from the interaction of ions with electrons and depends on the relative atomic weight, mass density and thickness of the sample according to Bethe-Bloch equation [8]. There are some drawbacks with common set-up/geometry of STIM imaging. In the case of multi-elemental sample having a variety mass density, the yield spectrum from each element has a different forward/ backward scattering kinematic factor then contributing to different energy loss. This will in turn decrease the contrast in the imaging [9]. As a another case, when focused MeV microbeams are scanned over a sample, the energy of the transmitted ions can be measured for each of the defined pixels and it can be used to form 2-D sample energy loss maps [10]. Although these images essentially can be formed from one ion in each pixel. To obtain images with more clarity, a number of counts per pixel are needed. It should be noted that these maps are not
representative of mass density due to the variation of energy loss caused by areal density variations of the sample. There are several ways in which stereoscopic images of ion microprobe can be used to obtain additional information on the sample. Previously, stereo-PIXE was used to quantitatively reconstruct the surface topography of engraved samples [11]. After that, stereoscopic PIXE imaging was displayed as anaglyphs using OMDAQ-3 software [12] to determine the three-dimensional distribution of the micrometeoroids impact in the form of craters from the Hubble Space Telescope [13]. Recently, dual-PIXE tomography system has been used to improve the X-ray detection to reconstruct Germanium in ICF target [14]. Although, stereoscopic STIM (stereo-STIM) imaging has been previously used to provide qualitative three-dimensional density information about certain specimens [15]. In this work, we follow a new approach with the introduction of a “disparity map” in stereo-STIM to generate a correspondence between points of the two images representing the same point of the sample in biological specimen. The use of stereo-STIM could also provide information about spatial distribution and orientation of the small features within a specimen. 2. Stereo-STIM model To study the effect of areal density variation on the result of analysis via stereo-STIM set-up a model of a thin sample with thickness of d is
E-mail address: [email protected]. https://doi.org/10.1016/j.nimb.2018.11.014 Received 11 October 2017; Received in revised form 28 October 2018; Accepted 9 November 2018 0168-583X/ © 2018 Elsevier B.V. All rights reserved.
Please cite this article as: Gholami Hatam, E., Nuclear Inst. and Methods in Physics Research B, https://doi.org/10.1016/j.nimb.2018.11.014
Nuclear Inst. and Methods in Physics Research B xxx (xxxx) xxx–xxx
E. Gholami Hatam
Fig. 1. A schematic layout of stereo-STIM model.
Fig. 2. The energy transmitted spectrum for different Mylar thicknesses. E1, E2 …E5 are the energies transmitted and are plotted by fitting the spectrum to a Gaussian distribution.
Fig. 3. (a) The energy difference versus the energy sum and (b) the disparity value for mean energies indicated at Fig. 2.
considered (Fig. 1). The energy E of a particle after passing through a layer of material with thickness x is given by the integral equation [16] where Ein is the initial energy
The relative areal density variation is embedded in this disparity map. As mentioned, the energy sum at denominator of disparity value is approximately constant for particles scattered at a certain depth while the energy difference in nominator is a fingerprint for inhomogeneity of the sample along the transmitted ions. Thus, true concentration maps can be created to resolve such imaging artefacts that are due to mass density variations.
x
E (x ) = Ein
(dE / dx ') dx '.
(1)
0
The energy sum of the right and left detectors in the case of scattering through 30° can be written as: x
(ER + EL)/2
E
K Ein
(dE / dx )0 dx 0
3. Experimental
3 (d x )/2 '
'
'
'
(dE / dx )1dx .
3.1. Experimental set-up
x
(2)
Using off-axis geometry makes it possible to have enough beam-currents that allow STIM to be used simultaneously with other ion beam analytical techniques such as PIXE and RBS. Stereo-STIM measurements were simultaneously performed with two off-axis STIM set-ups. The two identical particle detectors with resolution of 20 keV, 25 mm2 cross section and 40 mm detector-sample distance, were mounted on 30° and −30° with respect to the beam direction. The protons beam with spatial resolution lower than 10 μm and energy of 2 MeV were scanned over the samples area of 2.4 × 2.4 mm2 and the vacuum chamber pressure was maintained at 1.3 × 10−6 Torr at Van de Graaff accelerator in Tehran [17].
The stopping power, S = (dE/dx), before particles are scattered towards the detector is denoted with “0” and after scattering with “1”. The above equation could also be regarded as the mean of the energy, E , recorded in left and right detectors with respect to the beam direction. It has approximately constant value for particles scattered at depth x, since there is no preferred direction for particles to be scattered at the left or right with the same angle. The scattered protons at the depth of x travel the same distance through the sample, with the result that the relative difference in energy is zero for homogeneous samples; therefore, such events can be selected for further analysis. Accordingly, we have defined the “disparity value” that is implemented for each pixel in the left and right transmitted energy maps with this definition
(E ER ) D= L . (EL + ER )
3.2. Sample preparation For application of the new methodology presented in stereo-STIM an insect was chosen as a biological sample. It was glued to the part of the sample-holder that contains a hole to enhance the imaging and subsequently was air dried.
(3) 2
Nuclear Inst. and Methods in Physics Research B xxx (xxxx) xxx–xxx
E. Gholami Hatam
Fig. 4. (a) and (b) the energy transmitted map for whole body of a flying insect detected in stereo pair of left and right particle detectors. (c) The disparity map calculated from Eq. (3) for every pixel. (d), (e) and (f) are respectively the horizontal profiles of red line indicated in above maps. (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)
3.3. Normalization
i = 1, 2, …, 5. Here, Ē = E3 corresponds to Mylar with areal density of 35,000 (1015 atom/cm2). Fig. 3(b) presents the calculated disparity values of corresponding transmitted energies for known density variations. For discrepancies value close to zero, E ≈ Ē3, we do not have energy difference; hence, the generated disparity map would be null meaning that the sample is homogeneous. The lower (higher) disparity values indicate that the sample is thicker (thinner) than the proposed value of 35,000 (1015 atom/cm2). Interpolation must be used to implement the continuous function given discrete data points in order to determine the mass density variation function.
Regardless the nature of the sample, the value of disparity map in each pixel depends also on the statistic in summed spectra acquired from individual detectors. Even though two particle detectors have had the same geometry and resolution specification the different electronic adjustment may also affect the energy spectrum [18]. In order to consider this effect, a layer of 7 μm Kapton was used to normalize the recorded energy in each pixel of left and right maps. The value of the pixels in the individual energy map was balanced by the total number of particles detected at each detector for Kapton layer.
4.2. Biological sample
4. Result and discussion
As biological samples are mostly suitable to study with STIM analysis a small flying insect was scanned over its whole body. Fig. 4(a) and (b) represent the 2-D average transmitted energy recorded respectively at the left and right detectors. Just considering each of the individual map may ones mislead the interpretation of the result of STIM. Since according to the horizontal profile indicated in the centre of the (Fig. 4(d) and (e)) maps the abdomen part of the body in left detector has more counts with respect to the right spectrometer. At the same time, the head and eyes parts in the right detector have more counts with respect to the left spectrometer. Since the two particle detectors have the same geometry and the maps are normalised, these differences arise from the density variation or different topology along the beam direction that cannot be detected from a single detector spectrometer. Fig. 4(c) represents the disparity map for a whole region of the insect starting from the two point of views on it in the stereo pair. It is indicated from the horizontal profile in the disparity map (Fig. 4(f)) that the features of different parts of the body are resolved which are not seen in individual STIM data. The disparity map matches stereovision and takes the values between −1 (blue color) at the right detector where the left detector is blind and reaches the value of 1 (red color) at the left detector where the other detector is less sensitive.
4.1. Test sample At first, to test the performance of the proposed stereo-STIM model and also to calibrate the areal mass density, different thicknesses of Mylar foils were selected. The beam was scanned over the sample and the energy transmitted from 5 different foils were measured. The accurate mass densities were determined by fitting the corresponding spectra using SIMNRA software [19]. To present the transmitted energy in a two-dimensional map, the average or median energy value of the ions could be calculated in every pixel. Generally, average filtering is better in resolving small structures, even with sub-beam resolution for features of highly varying density [20]. This kind of filtering has been utilised in this study. Fig. 2 shows the spectrum for the transmitted energies corresponding to the 5 different thicknesses of Mylar. Fig. 3(a) illustrates how different mass densities can be resolved from a plot of the energy difference versus the energy sum. It is assumed that if the value of energies measured equals to E = (EL + ER )/2 = E3 and the E3 value is considered to be a reference for defining the change in areal mass densities. Then, the values of Ei − Ē versus Ei + Ē behave linear, where 3
Nuclear Inst. and Methods in Physics Research B xxx (xxxx) xxx–xxx
E. Gholami Hatam
5. Conclusion Application of procedures that can resolve small features within a specimen from a limited number of measurements or reconstruct specimens with non-uniform chemical composition will increase the range of specimens that can be analyzed. Since tomographic investigations are time-consuming and require very small sample dimensions, it could not replace stereo mapping delivering sufficient information in most applications. The stereo-STIM matches the two maps of the transmitted energy in the two detectors by introducing a disparity map. It was concluded that as long as the composition of the sample in known the disparity map is representative of the areal density variation. To validate the proposed model of stereo-STIM different thicknesses of Mylar were examined. In addition, an insect was also analyzed using the proposed stereo-STIM method to resolve different parts of its body. It was also deduced that removing a particular signal from one detector could have the effect of changing the precision of the target areal density as signaled by another detector. In summary, stereo-STIM can be a source of additional information simplifying the interpretation of the data and makes features visible, which cannot be separately distinguished in only two dimensions map.
[4] [5] [6] [7] [8] [9]
[10] [11] [12] [13]
Acknowledgement [14]
This research was supported by Malayer university, contact no. 84/ 9-1-452. I am also greatly indebted to Prof. M. Lamehi-Rachti from Nuclear Science Research School, Tehran, who provided insight and expertise that greatly assisted the research.
[15] [16]
References
[17]
[1] T. Andrea, M. Rothermel, T. Butz, T. Reinert, The improved STIM tomography setup at LIPSION: three-dimensional reconstruction of biological samples, Nucl. Instrum. Meth. Phys. Res. Sect. B 267 (2009) 2098–2102. [2] E.M. Stori, C.T. de Souza, L. Amaral, D. Fink, R.M. Papaléo, J.F. Dias, Use of STIM for morphological studies of microstructured polymer foils, Nucl. Instrum. Meth. Phys. Res. Sect. B 306 (2013) 99–103. [3] P. Aguer, L.C. Alves, P. Barberet, E. Gontier, S. Incerti, C. Michelet-Habchi, Z. Kertész, A.Z. Kiss, P. Moretto, J. Pallon, T. Pinheiro, J.E. Surlève-Bazeille,
[18] [19] [20]
4
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