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 Table of Contents  
Year : 2017  |  Volume : 8  |  Issue : 3  |  Page : 153-159

Synchrotron-based X-ray microimaging facility for biomedical research

Technical Physics Division, Bhabha Atomic Research Centre, Mumbai, Maharashtra, India

Date of Web Publication17-Oct-2017

Correspondence Address:
Ashish Kumar Agrawal
Technical Physics Division, Bhabha Atomic Research Centre, Mumbai, Maharashtra
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Source of Support: None, Conflict of Interest: None

DOI: 10.4103/jrcr.jrcr_29_17

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This paper reports the development of an X-ray imaging facility on Indus-2, synchrotron source at RRCAT, India and its potential applications in biomedical imaging. Indus-2 is country's first third-generation synchrotron source operating at energy of 2.5 Gev and current ~ 200 mA. It is a source of wide spectrum photon beam with very high flux and brilliance; therefore, it can be used for a variety of research such as spectroscopy, diffraction, imaging, lithography, and radiation processing. The development of X-ray imaging beamline here and implementation of several advanced imaging techniques, such as phase-contrast radiography, laminography, tomography, real-time imaging, and fluorescence imaging, has opened up new opportunities for characterization and study of soft tissue and biomaterials. This state of the art national facility is open to users for research in materials, biomedical and microtomography applications.

Keywords: Beamline, medical imaging, phase-contrast imaging, synchrotron, X-ray imaging

How to cite this article:
Agrawal AK, Singh B, Kashyap YS, Shukla M, Gadkari S C. Synchrotron-based X-ray microimaging facility for biomedical research. J Radiat Cancer Res 2017;8:153-9

How to cite this URL:
Agrawal AK, Singh B, Kashyap YS, Shukla M, Gadkari S C. Synchrotron-based X-ray microimaging facility for biomedical research. J Radiat Cancer Res [serial online] 2017 [cited 2023 Feb 5];8:153-9. Available from:

  Introduction Top

X-ray imaging has undergone revolutionary improvements, largely due to the availability of synchrotron sources and advanced detector technologies. This has opened up new possibilities in materials research and biomedical application.[1],[2],[3] The synchrotron source characteristics such as high brilliance, high coherence, and energy tunability have opened new horizons in imaging science such as phase-contrast imaging, diffraction-enhanced imaging (DEI), real-time imaging of transient phenomena, and in situ 3D imaging. High spatial Coherence and energy resolution at synchrotron facilities not only enables new modalities of imaging but also the conventional techniques such as tomography can be carried out with improved resolution. X-ray phase-contrast imaging has opened new possibilities to distinguish features with very small density difference. Therefore, this technique is very useful in biomedical research such as mammography, cancer research, food technology, evolutionary research, and study of biomaterials used in bones, teeth, and soft tissues. The possibility to use high-intensity monochromatic beam also allows element-specific imaging using multi-energy imaging, K-edge subtraction imaging,[4],[5],[6] etc.

With availability of Indus-2 synchrotron source at RRCAT, Indore, we have developed and installed an advanced X-ray imaging facility to carry out absorption and phase-sensitive imaging and microtomography for material and medical science application. The beamline has both monochromatic as well as white beam mode of operation. Several detectors such as charge-coupled device (CCD), flat panel, and an X-ray microscope with submicron resolution have been installed. Techniques such as radiography, propagation-based and diffraction-enhanced phase imaging, 3D tomography in absorption and phase-contrast mode, and real-time imaging have been implemented. This facility is being used for advanced applications in biomedical science research, nondestructive characterization of advanced materials and several new challenging applications.[4],[7],[8]

Basics of synchrotron-based advanced X-ray imaging techniques

Among all the other forms of radiation, X-rays have ability to penetrate into the bulk of the material. Due to smaller wavelengths, X-ray microscopy has a significantly higher spatial resolution than visible light microscopy. This allows for instance the study of biological specimen in water or the study of samples embedded in a solid. Moreover, X-ray-based imaging techniques are inherently nondestructive in nature, and no invasive sample preparation is necessary.[9],[10],[11] Hence, X-ray imaging has found wide application in the study of biological and technologically relevant materials. The behavior of X-rays as they travel through a sample can be described using a complex index of refraction, just as in conventional optics. In the X-ray region, the index of refraction, n, deviates only slightly from unity; it can be written as n = 1 – δ − iβ, where β describes the absorption of X-rays and δ the phase-shift term incorporates refractive effects. At typical x-ray energies of interest for soft material studies 15–100 keV, the phase-shift term can be up to 1000 times greater than the absorption term. Thus, it may be possible to observe phase contrast even if the absorption contrast is undetectable. Another factor, which favors phase contrast over absorption-based imaging, is the fact that phase term falls off less quickly. Phase-sensitive imaging delivers a complementary contrast with respect to absorption. To get information about the compounding of a sample, basically the density distribution of the sample, one has to relate the measured values for the refractive index to intrinsic parameters of the sample, such a relation is given by the followings: where ρais the atomic number density, p is phase-shift cross-section, σa is absorption cross-section, and κ is the X-ray wavenumber. Thus, the variation in the real part of refractive index is related to the phase-shift term while that in imaginary part is related to the attenuation of the object. While the attenuation of the X-rays can be measured with an X-ray detector, the phase shifts are not directly observable. Therefore, an optical system is generally needed to convert the phase shift into intensity modulations. Depending on the method of converting this phase information into intensity modulation, over the year, a plethora of the techniques have been developed [Figure 1]. They can be grouped into following categories: (1) Free space propagation-based imaging (PBI), (2) DEI, (3) Grating-based differential phase-contrast imaging (grating-based imaging), and (4) Coded aperture-based imaging.[5],[12],[13],[14] These techniques differ in their complexity of instrumental implementation or quantitative analysis for extracting information out of acquired images. The phase-sensitive techniques rely on high spatial and temporal coherence of X-ray beams, which are available only at synchrotron sources. However, some of them such as free space PBI, coded aperture-based imaging, and grating-based differential phase- contrast imaging, which only need high spatial coherence, can be carried out using polychromatic microfocus sources.
Figure 1: Different variants of X-ray phase-contrast imaging (a) Bonse–Hart interferometer (b) diffraction-enhanced imaging (c) grating-based imaging (d) propagation-based imaging

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Synchrotron-based X-ray imaging beamline

We have developed a state of the art imaging facility dedicated to biomedical and material science research at Indus-2 synchrotron source operating at 2.5 Gev @300 mA. It has provision to use both monochromatic as well as polychromatic (white) X-rays. Monochromatic beam is required for dual-energy imaging, diffraction-enhanced imaging, and improved phase-contrast imaging. On the other hand, white beam with higher flux and wider energy range is required for real-time imaging and imaging of thicker samples. The energy range covered by this beamline is 8–35 keV in monochromatic mode and up to 60 keV in white beam mode. Absorption as well as phase imaging techniques has been implemented in the same experimental hutch. The new imaging techniques exploit the coherence property of the X-ray source and provide improved contrast. Typical design parameters and available imaging instruments are listed in [Table 1].
Table 1: Design parameters of imaging beamline

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The beamline comprises optics hutch, experimental station, control room, and data analysis facility.[2] Optics hutch comprises components for beam shape and size selection, monochromatization, beam filtering, exposure and dose minimization, etc., such as Water-cooled Be window assembly, vertical and horizontal entrance slits, double crystal monochromator (DCM), vertical and horizontal exit slits, beam shutter and beam position monitor, and water-cooled exit Be window assembly for monochromatic and white beam are installed in optics hutch. [Figure 2] shows the optical layout of the imaging beamline with details of various instruments included along with their distances from the source.
Figure 2: Optical layout of the imaging beamline

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The experimental station consists of all the instruments required for image formation, sample and detector manipulation, sample mounting, sample environments for in situ imaging, optical instruments for image formation and analysis, etc., [Figure 3]. Following detectors are available as per the experimental requirements:
Figure 3: Photograph of the experimental station of imaging beamline

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  1. X-ray imaging microscope - a unique facility to enable submicrometer-resolution imaging (best achievable resolution is 700 nm). It consists of a lens-coupled CCD detector with mirror optics and variety of combinations of scintillators and objectives for choosing required resolution (700 nm - 12.8 um), field of view (0.76 mm - 12 mm)
  2. High-resolution X-ray CCD detector - fiber-optic coupled, Gadox scintillator with 4007 × 2678 (18 mm × 12 mm), pixel size 4.5μm
  3. X-ray flat panel detector - 120 mm field of view, 50 μm pixel size, 2400 × 2400 pixels
  4. Image plate detectors – high-sensitivity imaging for large sample (readout resolution up to 10 μm)

To align sample and detectors with respect to incident beam following set of sample and detector manipulation, stages are used:

  1. High precision six-axis sample manipulator stages consisting of Y, Z, θ, ψ, ϕ
  2. High precision three-axis manipulator for detectors consisting X, Y, Z motions.

Following other instruments are also available for the implementation of advanced imaging techniques and facilitation of various experimental requirements:

  1. Ion chamber for online beam current measurement and dose regulation
  2. Fast shutter for controlled exposure time in biomedical imaging
  3. Analyzer DCM with special mount to perform phase-contrast imaging, diffraction-enhanced imaging/ultra-small angle scattering experiments
  4. 3kN and 500N load cell assembly for in situ experiments under load conditions in white beam mode only
  5. Complete experimental station on vibration isolated granite tables.

Experimental techniques at imaging beamline

Imaging beamline is dedicated for X-ray radiography and tomography applications in the field of materials and medical science. Radiography and tomography in absorption contrast are useful in obtaining the distribution of microstructural features, density, and the attenuation coefficients in 2D and 3D. Spatial distribution of enclosed phases, their morphology, architecture or geometry, imperfections such as voids, crack, and porosity can be visualized and quantified. Propagation-based phase-contrast imaging for weakly absorbing materials can be used to obtain the refractive index distribution. Dual-energy imaging at absorption edges is useful for mapping the spatial distribution of specific elements.[15],[16] A brief description of all the available techniques at imaging beamline and their applicability of given in [Table 2].
Table 2: Various techniques at imaging beamline and their brief description

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Biomedical application of imaging beamline

With the implementation of various phase-contrast X-ray imaging techniques, sensitivity to small density variations at microscopic scale is tremendously improved as compared to conventional imaging; therefore, experimental setup at imaging beamline is most suitable for microimaging of soft materials, biological tissues, small animal samples, and biomaterials for various biomedical research objectives. Extensive review articles have been published in the literature to discuss the wide range of biomedical applications of synchrotron-based X-ray imaging techniques and their merits over conventional imaging.[9],[12],[14],[17],[18],[19],[20] In diagnostic imaging, image-based characterization is helpful in disease diagnostics, identification of root cause, location of abnormalities of structure, or function. Preclinical study for various diagnostic imaging such as mammography, functional lung imaging, and bone imaging is active fields of research where X-ray phase-contrast imaging is playing a very active role. Similarly, in developmental biology research, studies on evolution and growth of various plants and animals by comparing the morphological and anatomical features at various stages of development are very helpful in understanding the effects of various nutritional, toxicological, and environmental factors. Studies to determine the effects of various drug treatments on animal and plant tissues and organ can also be carried out. X-ray microimaging techniques are helpful in optimizing the microstructure of biomaterials such as bone scaffolds to achieve desired mechanical, transport, and biocompatible properties.

  Results and Discussion Top

A large variety of experiments have been conducted at imaging beamline to address various research problems in materials science, biomedical science, clinical imaging, agriculture, paleontology, geology, environmental science, engineering, and dosimetry. It is impossible to include all the results in the single report, and we give an overview of some of the example studies carried out on various samples.

Cancer research

Mammography is a popular technique for breast cancer screening and low-dose diagnostic imaging. CIRS tissue-equivalent phantom is the standard used to assess the capability and threshold of the various mammography machines. To study the feasibility of mammography and cancerous lesions detection at imaging beamline, we have used this phantom and imaged various targets in it which simulate calcifications, fibrous calcifications in ducts, and tumor masses. [Figure 4] shows the phase-contrast image of representative microcalcifications of group 8. This group represents CaCO3 specs with 230 μ grain size. The phase-contrast images acquired can clearly identify the microcalcifications in this tissue-equivalent phantom along with their respective shapes and sizes. The experiment was carried out at 25 keV, and sample to detector distance was optimized at 300 mm. The experiment clearly demonstrates the utility of experimental setup for tumor detection and cancer research on small animals and biopsy samples.[21],[22],[23]
Figure 4: (a) Tissue-equivalent mammography phantom (b) phase-contrast image of CaCo3 specs in group 8 with 230 μ grain size

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Anatomy of small animals or insects

To show the utility of synchrotron-based phase-contrast imaging in small animals anatomy and internal organ nondestructively, we have taken the example of zebrafish and house bee as their representatives. Propagation-based phase-contrast image of zebrafish is shown in [Figure 5] and its internal organs such as bones, head, glands, and feathers can be easily seen. Various studies related to chemical or environmental toxicity and drug treatment are possible to see their effects on organ anatomy and morphology.In vivo imaging of small animals and insects also permits us to see functioning of various organs such as lungs and heart. [Figure 6] shows the diffraction-enhanced phase-contrast image of house bee. Very small and low absorbing anatomical features such as wings are also clearly seen in this image. This can be attributed to very high sensitivity of diffraction-enhanced imaging technique.[21],[24],[25] Because of requirement of highly precise instrumentation and requirement of monochromatic source, this technique is available only at synchrotron source-based imaging facilities.
Figure 5: Propagation-.based phase-.contrast image of zebrafish showing anatomy of internal organs

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Figure 6: Diffraction-enhanced image house bee showing anatomy of various organs

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Microstructures of plants tissues and organs

As a representative of this, we have taken leaf, stem, and seeds of soyabean. The studies carried out on these plant organs using phase-contrast imaging and microtomography revealed their respective microstructures. In case of leaves, microstructure of midrib and much finer vein structure of the 2nd, 3rd, and higher order [Figure 7] can be seen clearly. Apart from this, the image also shows the spatial distribution of oxalate crystals and their localized deposition at places. With this feasibility study, several experiments have been conducted to study the effects of ultraviolet filtering under different bands on the plant leaf microstructure and thereby on plants overall growth.[26],[27],[28] Similarly, effect of treatment with magnetic field of different strength to the seed before sowing was also studied on lead vein structure and thereby on plant growth using quantitative phase-contrast imaging. Other study was also done to see the effect of fungicide treatment on plant leaves, etc., In case of seed, the microtomography studies were carried out to see process of their germination. As shown in [Figure 8], the microtomography images clearly show the microstructure of seed and germination point.
Figure 7: Phase-contrast microimaging of soyabean plant leaf to see its vascular microstructure and deposited oxalate crystals

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Figure 8: Microtomographic imaging of soyabean plant seeds and stems to see their microstructure. (a and b) Microtomography reconstructed slice images of soyabean seed (c and d) 3D image of soyabean seed (e and f) microtomography reconstructed slice images of soyabean plant stem (g and h) 3D image of soyabean plant stem

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As a representative of this, we have carried out experiments on human teeth to study the root canal morphology. As shown in [Figure 9], the microtomography images clearly distinguish structure of dentin, enamel, and root canal in reconstructed slices and 3D images. Using quantitative analysis, shape, size, and orientation can also be measured for these features. With this feasibility experiment on teeth, we have carried out a study to find quality of restoration to broken human teeth using various composite resins.[29] The study was conducted on various teeth types, and tooth-resin interface as well as porosity/imperfections of restored region were visualized and quantified to evaluate the quality of restoration.
Figure 9: Microtomographic imaging of human teeth to see its root canal microanatomy and quality of restoration when restored with composite resins. (a) Micro-CT reconstructed slice image of human tooth (b) 3D volume image showing tooth internal structure (c) Interface of broken tooth with restored part (d) porosity in the restored region

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  Conclusions Top

Synchrotron-based X-ray microimaging facility at Indus-2 synchrotron source is a unique facility within country for biomedical research. It has advanced imaging modes such as phase-contrast imaging, tomography, and diffraction-enhanced imaging, and the facility is capable of providing unmatched X-ray image quality and sensitivity. The experimental setup along with brief description of various imaging modes is discussed to show the potential of the facility and its applicability for variety of biomedical research problems.[30]

Financial support and sponsorship


Conflicts of interest

There are no conflicts of interest.

  References Top

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Available from: [Last accessed on 2017 Aug 31].  Back to cited text no. 30


  [Figure 1], [Figure 2], [Figure 3], [Figure 4], [Figure 5], [Figure 6], [Figure 7], [Figure 8], [Figure 9]

  [Table 1], [Table 2]

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