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Despite the current advances in micro-CT analysis, the influence of some image acquisition parameters on the morphometric assessment outcome have not been fully elucidated. The aim of this study was to determine whether data binning and frame averaging affect the morphometric outcome of bone repair assessment using micro-CT. Four Wistar rats' tibiae with a surgically created bone defect were imaged with micro-CT six times each, frame averaging set to 1 and 2, and data binning set to 1, 2 and 4, for each of the averaging values. Two-way ANOVA followed by Bonferroni tests assessed the significance of frame averaging and data binning on a set of morphometric parameters assessed in the image volumes (p < 0.01). The effect of frame averaging was not significant for any of the assessed parameters. Increased data binning led to larger trabecular thickness. In contrast, smaller bone volume fraction and bone volume were found as data binning increased. Trabeculae number and trabecular separation were not influenced by any of the parameters. In conclusion, the morphometric outcome of bone repair assessment in micro-CT demonstrated dependency upon data binning, but not frame averaging. Therefore, image acquisition of small anatomical structures (e.g., rat trabeculae) should be performed without data binning., (© 2022. The Author(s).)

Micro-CT provides critical data for musculoskeletal research, yielding three-dimensional datasets containing distributions of mineral density. Using high-resolution scans, we quantified changes in the fine architecture of bone in the spine of young mice. This data is made available as a reference to physiological cancellous bone growth. The scans (n = 19) depict the extensive structural changes typical for female C57BL/6 mice pups, aged 1-, 3-, 7-, 10- and 14-days post-partum, as they attain the mature geometry. We reveal the micro-morphology down to individual trabeculae in the spine that follow phases of mineral-tissue rearrangement in the growing lumbar vertebra on a micrometer length scale. Phantom data is provided to facilitate mineral density calibration. Conventional histomorphometry matched with our micro-CT data on selected samples confirms the validity and accuracy of our 3D scans. The data may thus serve as a reference for modeling normal bone growth and can be used to benchmark other experiments assessing the effects of biomaterials, tissue growth, healing, and regeneration.

Mitochondria are dynamic organelles that change morphology to adapt to cellular energetic demands under both physiological and stress conditions. Cardiomyopathies and neuronal disorders are associated with structure-related dysfunction in mitochondria, but three-dimensional characterizations of the organelles are still lacking. In this study, we combined high-resolution imaging and 3D electron density information provided by cryo-soft X-ray tomography to characterize mitochondria cristae morphology isolated from murine. Using the linear attenuation coefficient, the mitochondria were identified (0.247 ± 0.04 µm -1 ) presenting average dimensions of 0.90 ± 0.20 µm in length and 0.63 ± 0.12 µm in width. The internal mitochondria structure was successfully identified by reaching up the limit of spatial resolution of 35 nm. The internal mitochondrial membranes invagination (cristae) complexity was calculated by the mitochondrial complexity index (MCI) providing quantitative and morphological information of mitochondria larger than 0.90 mm in length. The segmentation to visualize the cristae invaginations into the mitochondrial matrix was possible in mitochondria with MCI ≥ 7. Altogether, we demonstrated that the MCI is a valuable quantitative morphological parameter to evaluate cristae modelling and can be applied to compare healthy and disease state associated to mitochondria morphology.

Tracheal tissue engineering has become an active area of interest among clinical and scientific communities; however, methods to evaluate success of in vivo tissue-engineered solutions remain primarily qualitative. These evaluation methods have generally relied on the use of photographs to qualitatively demonstrate tracheal patency, endoscopy to image healing over time, and histology to determine the quality of the regenerated extracellular matrix. Although those generally qualitative methods are valuable, they alone may be insufficient. Therefore, to quantitatively assess tracheal regeneration, we recommend the inclusion of microcomputed tomography (μCT) to quantify tracheal patency as a standard outcome analysis. To establish a standard of practice for quantitative μCT assessment for tracheal tissue engineering, we recommend selecting a constant length to quantify airway volume. Dividing airway volumes by a constant length provides an average cross-sectional area for comparing groups. We caution against selecting a length that is unjustifiably large, which may result in artificially inflating the average cross-sectional area and thereby diminishing the ability to detect actual differences between a test group and a healthy control. Therefore, we recommend selecting a length for μCT assessment that corresponds to the length of the defect region. We further recommend quantifying the minimum cross-sectional area, which does not depend on the length, but has functional implications for breathing. We present empirical data to elucidate the rationale for these recommendations. These empirical data may at first glance appear as expected and unsurprising. However, these standard methods for performing μCT and presentation of results do not yet exist in the literature, and are necessary to improve reporting within the field. Quantitative analyses will better enable comparisons between future publications within the tracheal tissue engineering community and empower a more rigorous assessment of results. Impact statement The current study argues for the standardization of microcomputed tomography (μCT) as a quantitative method for evaluating tracheal tissue-engineered solutions in vivo or ex vivo. The field of tracheal tissue engineering has generally relied on the use of qualitative methods for determining tracheal patency. A standardized quantitative evaluation method currently does not exist. The standardization of μCT for evaluation of in vivo studies would enable a more robust characterization and allow comparisons between groups within the field. The impact of standardized methods within the tracheal tissue engineering field as presented in the current study would greatly improve the quality of published work.

Background: Micro-computed tomography (CT) is a non-invasive technique that is used to obtain three-dimensional (3D) images of tissue structure in small animals. Compared with extensive mammal studies, few 3D imaging studies of fish have been conducted using micro-CT. An optimized method for imaging fish tissue structure is necessary, because they have adapted to diverse environments via functional and structural specialization., New Method: Brains of three species with different sizes and habitats were fixed in 4% paraformaldehyde and immersed in non-ionic iodinated contrast agent (Iopamiron). We examined the relationship between Iopamiron concentration and immersion time to determine universally optimal conditions for use in fish., Results: We reconstructed 3D images of whole fish brains from cross-sections of brains from the Malabar grouper (Epinephelus malabaricus), bastard halibut (Paralichthys olivaceus), and threespot wrasse (Halichoeres trimaculatus). Developmental changes in brain structure were observed in the bastard halibut. Most brain regions of the threespot wrasse were distinguishable, although inner regions of the brain were less visible., Comparison With Existing Methods: Histological techniques are typically used to observe fish brain structure, despite its drawbacks in terms of tissue sample preparation (shrinkage and distortion) and image capture (3D image constriction). The technique examined in the present study solves these problems and allows for the simultaneous handling of multiple specimens., Conclusion: Micro-CT imaging is suitable for observing the surfaces and inner structures of fish of various species., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Digital Volume Correlation (DVC) is used to measure internal displacements and strains in bone. Recent studies have shown that Synchrotron radiation micro-computed tomography (SR-microCT) can improve the accuracy and precision of DVC. However, only zero-strain or virtually-moved test have been used to quantify the DVC uncertainties, leading to potential underestimation of the measurement errors. In this study, for the first time, the uncertainties of a global DVC approach have been evaluated on repeated SR-microCT scans of bovine cortical bone (voxel size: 1.6 μm), which were virtually deformed for different magnitudes and along different directions. The results showed that systematic and random errors of the normal strain components along the deformation direction were higher than the errors along unstrained directions. The systematic percentage errors were smaller for larger virtual deformations. The random percentage error was in the order of 10% of the virtual deformation. However, higher errors were localized at the boundary of the volumes of interest, perpendicular to the deformation direction. When only the central region of the samples was considered (100 µm layers removed from the borders where the deformation was applied), the errors in the direction of virtual deformation were comparable to the errors in the unstrained directions. In conclusion, the method presented to estimate the uncertainties of DVC is suitable for testing anisotropic specimens as cortical bone. The good agreement between the uncertainties in measurements of strain components obtained with this approach and with the simpler zero-strain-test suggests that the latter is adequate in the tested deformation scenarios., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Background: Ultrasonography is a fast and patient-friendly modality to assess cartilage thickness. However, inconsistent results regarding accuracy have been reported. Therefore, we asked what are (1) the accuracy, (2) reproducibility, and (3) reliability of ultrasonographic cartilage thickness measurement using contrast-enhanced micro-CT for validation?, Methods: A series of 50 cartilage-bone plugs were harvested from fresh bovine and porcine joints. Ultrasonic cartilage thickness was determined using an A-mode, 20-MHz hand-held ultrasonic probe with native (1580 m/s) and adjusted speed of sound (1696 m/s). All measurements were performed by two observers at two different occasions. Angle of insonation was controlled by tilting the device and recording minimal thickness. Retrieval of exact location for measurement was facilitated by aligning the circular design of both cartilage-bone plug and ultrasonic device. There was no soft tissue interference between cartilage surface and ultrasonic probe. Ground truth measurement was performed using micro-CT with iodine contrast agent and a voxel size of 16 μm. The mean cartilage thickness was 1.383 ± 0.402 mm (range, 0.588-2.460 mm)., Results: Mean accuracy was 0.074 ± 0.061 mm (0.002-0.256 mm) for native and 0.093 ± 0.098 mm (0.000-0.401 mm) for adjusted speed of sound. Bland-Altman analysis showed no systematic error. High correlation was found for native and adjusted speed of sound with contrast-enhanced micro-CT (both r = 0.973; p < 0.001). A perfect agreement for reproducibility (intraclass correlation coefficient [ICC] 0.992 and 0.994) and reliability (ICC 0.993, 95% confidence interval 0.990-0.995) was found., Conclusions: Ultrasonic cartilage thickness measurement could be shown to be highly accurate, reliable, and reproducible. The A-mode ultrasonic cartilage thickness measurement is a fast and patient-friendly modality which can detect early joint degeneration and facilitate decision making in joint preserving surgery.

Objective: The aim of this paper was to develop calibration standards (CSs) that are readily available for clinical researchers for the quantitation of enamel mineral content., Method: Polyethylene terephthalate (PET), acetal, polyphenylene sulfide (PPS), selenite, Egyptian alabaster, aragonite, and fluorite were fashioned into discs, and their densities were measured and stacked for microcomputed tomography examination. Frame averaging, flat-field correction, pre-filtration, and beam-hardening correction were applied. CSs were checked for homogeneity. The linear relationship between the mean greyscale value (GSV) of each disc and its physically calculated density was explored, and reproducibility was tested. A calibration function was established and then validated using a bovine enamel disc and sound enamel of extracted human premolar teeth., Results: Measured densities were PET (ρ = 1.38 g/cm3), acetal (ρ = 1.41 g/cm3), PPS (ρ = 1.64 g/cm3), selenite (ρ = 2.24 g/cm3), Egyptian alabaster (ρ = 2.7 g/cm3), aragonite (ρ = 2.72 g/cm3), and fluorite (ρ = 3.11 g/cm3). Examination of the profile sections of CSs confirmed the uniformity of GSVs with minimal beam-hardening effect. A squared Pearson correlation coefficient of R2 = 0.994 was determined between the mean GSV of each CS and its calculated density and was reproduced at different settings with R2 >0.99. A linear regression equation of density (y) versus GSV (x) was established using the least squares regression equation method. The estimated density of the bovine enamel disc (2.48 g/cm3) showed high accuracy when compared to the physically measured value (2.45 g/cm3). The -relative error was 1.2%. Densities of sound enamel in the extracted human premolar teeth were 2.6-3.1 g/cm3., Conclusions: This is a simple, valid, and reproducible method to quantitate enamel mineral content. This simple, yet accurate system could be used to expand knowledge in the field of enamel caries research., (©2019 The Author(s) Published by S. Karger AG, Basel.)

While several materials and techniques have been used to assess the quality of root canal fillings in micro-CT images, the lack of standardization in scanning protocols has produced conflicting results. Hence, the aim of this study was to determine a cutoff voxel size value for the assessment of root canal filling voids in micro-CT and nano-CT images. Twenty freshly extracted mandibular central incisors were used. Root canals were prepared with nickel titanium files to an ISO size 40/0.06 taper and then filled with a single cone (40/0.06 taper) and AH Plus sealer. The teeth were scanned with different voxel sizes with either micro-CT (5.2, 8.1, 11.2, and 16.73  μ m) or nano-CT (1.5 and 5.0  μ m) equipment. Images were reconstructed and analyzed with the NRecon and CTAn software. Void proportion and void volume were calculated for each tooth in the apical, middle, and coronal thirds of the root canal. Kruskal-Wallis and post hoc Mann-Whitney U tests were performed with a significance level of 5%. In micro-CT images, significantly different results were detected among the tested voxel sizes for void proportion and void volume, whereas no such differences were found in nano-CT images ( p > 0.05). Micro-CT images showed higher void numbers over the entire root length, with statistically significant differences between the voxel size of 16.73  μ m and the other sizes ( p < 0.05). The values of the different nano-CT voxel sizes did not significantly differ from those of the micro-CT (5.2, 8.1, and 11.2  μ m), except for the voxel size of 16.73  μ m ( p < 0.05). All tested voxel sizes enabled the detection of root canal filling voids except for the voxel size of 16.73  μ m. Bearing in mind the limitations of this study, it seems that a voxel size of 11.2  μ m can be used as a reliable cutoff value for the assessment of root canal filling voids in micro-CT imaging.

The goal of this work was to establish imaging dose to mouse organs with a validated Monte Carlo (MC) model of the image-guided Small Animal Radiation Research Platform (SARRP) and to investigate the effect of scatter from the internal walls on animal therapy dose determination. A MC model of the SARRP was built in the BEAMnrc code and validated with a series of homogeneous and heterogeneous phantom measurements. A segmented microCT scan of a mouse was used in DOSXYZnrc to determine mouse organ microCT imaging doses to 15-35 g mice for the SARRP pancake (mouse lying on couch) and standard (mouse standing on couch) imaging geometries for 40-80 kVp tube voltages. Imaging dose for off-center positioning shifts and maintaining image noise across tube voltages were also calculated. Half-value layer (HVL) measurements for the 220 kVp therapy beam in the presence of the SARRP shielding cabinet were modeled in BEAMnrc and compared to the 100 cm source-to-detector distance (SDD) in the scatter free, narrow-beam geometry recommended by the American Association of Physicists in Medicine Task Group 61 (AAPM TG-61). For a 60 kVp, 0.8 mA, and 60 s scan protocol, maximum mean organ imaging doses to boney and non-boney structures were 10.5 cGy and 3.5 cGy, respectively, for an average size 20 g mouse. Current-exposure combinations above 323, 203, 147, 116, and 95 mAs for 40-80 kVp tube voltages, respectively, will increase body doses above 10 cGy. MicroCT mean body dose was 18% lower in pancake compared to standard imaging geometry. An 11% difference in measured HVL at a 50 cm SDD was found compared to MC simulated HVL for the AAPM TG-61 recommended scatter free geometry at a 100 cm SDD. This change in HVL resulted in a 0.5% change in absorbed dose to water calculations for the treatment beam.

Background: Quantitative assessment of post-transcatheter aortic valve replacement (TAVR) aortic regurgitation (AR) remains challenging. We developed patient-specific anatomical models with pulsatile flow circuit and investigated factors associated with AR after TAVR., Methods: Based on pre-procedural computed tomography (CT) data of the six patients who underwent transfemoral TAVR using a 23-mm SAPIEN XT, anatomically and mechanically equivalent aortic valve models were developed. Forward flow and heart rate of each patient in two days after TAVR were duplicated under mean aortic pressure of 80mmHg. Paravalvular leakage (PVL) volume in basal and additional conditions was measured for each model using an electromagnetic flow sensor. Incompletely apposed tract between the transcatheter and aortic valves was examined using a micro-CT., Results: PVL volume in each patient-specific model was consistent with each patient's PVL grade, and was affected by hemodynamic conditions. PVL and total regurgitation volume increased with the mean aortic pressure, whereas closing volume did not change. In contrast, closing volume increased proportionately with heart rate, but PVL did not change. The minimal cross-sectional gap had a positive correlation with the PVL volumes (r=0.89, P=0.02). The gap areas typically occurred in the vicinity of the bulky calcified nodules under the native commissure., Conclusions: PVL volume, which could be affected by hemodynamic conditions, was significantly associated with the minimal cross-sectional gap area between the aortic annulus and the stent frame. These data may improve our understanding of the mechanism of the occurrence of post-TAVR PVL., (Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.)

Selecting identical topographical locations to analyse pathological structural changes of the osteochondral unit in translational models remains difficult. The specific aim of the study was to provide objectively defined reference points on the ovine tibial plateau based on 2-D sections of micro-CT images useful for reproducible sample harvesting and as standardized landmarks for landmark-based 3-D image registration. We propose 5 reference points, 11 reference lines and 12 subregions that are detectable macroscopically and on 2-D micro-CT sections. Their value was confirmed applying landmark-based rigid and affine 3-D registration methods. Intra- and interobserver comparison showed high reliabilities, and constant positions (standard errors < 1%). Spatial patterns of the thicknesses of the articular cartilage and subchondral bone plate were revealed by measurements in 96 individual points of the tibial plateau. As a case study, pathological phenomena 6 months following OA induction in vivo such as osteophytes and areas of OA development were mapped to the individual subregions. These new reference points and subregions are directly identifiable on tibial plateau specimens or macroscopic images, enabling a precise topographical location of pathological structural changes of the osteochondral unit in both 2-D and 3-D subspaces in a region-appropriate fashion relevant for translational investigations.

Microfocus X-ray computed tomography (micro-CT) has been applied as a method for the nondestructive and detailed assessment of trabecular bone patterns and tooth structure. Voxel values obtained from micro-CT are not absolute values. Therefore, voxel values were assessed using hydroxyapatite (HA) blocks with a different vesicle rate to quantify voxel values of micro-CT images in the present investigation.HA blocks with 4 levels of porosity and a block with a soft tissue-equivalent density were used, and the voxel values of each block were measured. Correlations between voxel values of micro-CT and HA densities were analyzed. Also, black and white binary images were produced, and the ratios of white pixels to pixels in regions of interest (ROIs) were calculated. The relationship between voxel values of micro-CT and HA densities could be regressed using a linear equation, and the correlation coefficient was high. Also, there were no significant differences in the regression equations between the first and second times. Voxel values of micro-CT might be convertible to HA densities using a regression equation.

In this study, we assess the precision, accuracy, and repeatability of craniodental landmarks (Types I, II, and III, plus curves of semilandmarks) on a single macaque cranium digitally reconstructed with three different surface scanners and a microCT scanner. Nine researchers with varying degrees of osteological and geometric morphometric knowledge landmarked ten iterations of each scan (40 total) to test the effects of scan quality, researcher experience, and landmark type on levels of intra- and interobserver error. Two researchers additionally landmarked ten specimens from seven different macaque species using the same landmark protocol to test the effects of the previously listed variables relative to species-level morphological differences (i.e., observer variance versus real biological variance). Error rates within and among researchers by scan type were calculated to determine whether or not data collected by different individuals or on different digitally rendered crania are consistent enough to be used in a single dataset. Results indicate that scan type does not impact rate of intra- or interobserver error. Interobserver error is far greater than intraobserver error among all individuals, and is similar in variance to that found among different macaque species. Additionally, experience with osteology and morphometrics both positively contribute to precision in multiple landmarking sessions, even where less experienced researchers have been trained in point acquisition. Individual training increases precision (although not necessarily accuracy), and is highly recommended in any situation where multiple researchers will be collecting data for a single project.

Micro computed tomography (micro-CT) has already been proposed as a useful technique for the qualitative analysis of false starts (FS) produced on human bones, although the reliability and the error rate of this technique have not been tested yet, neither for qualitative nor for quantitative assessments. The aim of the present study was to test the morphological agreement, accuracy, precision and inter-rater reliability of micro-CT analysis of FS on bones. The morphological agreement was assessed through the degree of concordance among the 3 independent blind raters in the identification of the shape of 24 FS manually produced on bones by 3 different saws (8 FS for each saw). The accuracy was calculated through the percentage of error in the automatic and manual measurement of the diameter of a reference object. The precision was calculated as CV% of multiple measurements performed by 3 independent blind raters on the reference object and one bone sample acquired 20 times. The inter-rater reliability was assessed as intraclass correlation coefficient (ICC) among measurements performed by 3 independent blind raters, assessing 24 FS produced using 3 different saws. The results demonstrated that both qualitative and quantitative analysis were reproducible and robust. Micro-CT analysis showed a 100% morphological agreement, a high level of accuracy (percentage error < 0,5%), precision (CV% < 5%) and inter-rater reliability (ICC > 0.995), when FS were analyzed by forensic pathologists and/or radiologists with adequate expertise. Obviously, further validation studies are needed, including a higher number of samples produced by a wider variety of saws and multiple operators., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Sometimes, drug product for parenteral administration is stored in a frozen state (e.g., -20 °C or -80 °C), particularly during early stages of development of some biotech molecules in order to provide sufficient stability. Shipment of frozen product could potentially be performed in the frozen state, yet possibly at different temperatures, for example, using dry ice (-80 °C). Container closure systems of drug products usually consist of a glass vial, rubber stopper, and an aluminum crimped cap. In the frozen state, the glass transition temperature (Tg) of commonly used rubber stoppers is between -55 and -65 °C. Below their Tg, rubber stoppers are known to lose their elastic properties and become brittle, and thus potentially fail to maintain container closure integrity in the frozen state. Leaks during frozen temperature storage and transportation are likely to be transient, yet, can possibly risk container closure integrity and lead to microbial contamination. After thawing, the rubber stopper is supposed to re-seal the container closure system. Given the transient nature of the possible impact on container closure integrity in the frozen state, typical container closure integrity testing methods (used at room temperature conditions) are unable to evaluate and thus confirm container closure integrity in the frozen state. Here we present the development of a novel method (thermal physical container closure integrity) for direct assessment of container closure integrity by a physical method (physical container closure integrity) at frozen conditions, using a modified He leakage test. In this study, different container closure systems were evaluated with regard to physical container closure integrity in the frozen state to assess the suitability of vial/stopper combinations and were compared to a gas headspace method. In summary, the thermal physical container closure integrity He leakage method was more sensitive in detecting physical container closure integrity impact than gas headspace and aided identification of an unsuitable container closure system., Lay Abstract: Sometimes, drug product for parenteral administration is stored in a frozen state (e.g., -20 °C or -80 °C), particularly during early stages of development of some biotech molecules in order to provide sufficient stability. Container closure systems for drug products usually consist of a glass vial, rubber stopper, and an aluminum crimped cap. In the frozen state, the glass transition temperature (Tg) of commonly used rubber stoppers is between -55 and -65 °C. Leaks during frozen temperature storage and transportation are likely to be transient, yet they can possibly risk container closure integrity and lead to microbial contamination and sterility breach. After thawing, the rubber stopper is expected to re-seal the container closure system. Given the transient nature of the possible impact on container closure integrity in the frozen state, typical container closure integrity testing methods (used at room temperature conditions) are unable to evaluate and thus confirm container closure integrity in the frozen state. Here we present the development of a novel method (thermal container closure integrity) for direct measurement of container closure integrity by a physical method (physical container closure integrity) at frozen conditions, using a modified He leakage test. In this study, we found that the thermal container closure integrity He leakage method was more sensitive in detecting physical container closure integrity impact than gas headspace and aided identification of an unsuitable container closure system., (© PDA, Inc. 2016.)

Endplate morphology is understood to play an important role in the mechanical behavior of vertebral bone as well as degenerative processes in spinal tissues; however, the utility of clinical imaging modalities in assessment of the vertebral endplate has been limited. The objective of this study was to evaluate the ability of two clinical imaging modalities (digital tomosynthesis, DTS; high resolution computed tomography, HRCT) to assess endplate topography by correlating the measurements to a microcomputed tomography (μCT) standard. DTS, HRCT, and μCT images of 117 cadaveric thoracolumbar vertebrae (T10-L1; 23 male, 19 female; ages 36-100 years) were segmented, and inferior and superior endplate surface topographical distribution parameters were calculated. Both DTS and HRCT showed statistically significant correlations with μCT approaching a moderate level of correlation at the superior endplate for all measured parameters (R(2)Adj=0.19-0.57), including averages, variability, and higher order statistical moments. Correlation of average depths at the inferior endplate was comparable to the superior case for both DTS and HRCT (R(2)Adj=0.14-0.51), while correlations became weak or nonsignificant for higher moments of the topography distribution. DTS was able to capture variations in the endplate topography to a slightly better extent than HRCT, and taken together with the higher speed and lower radiation cost of DTS than HRCT, DTS appears preferable for endplate measurements., (Copyright © 2015 Elsevier Inc. All rights reserved.)

The quality of Computed Tomography (CT) images crucially depends on the precise knowledge of the scanner geometry. Therefore, it is necessary to estimate and calibrate the misalignments before image acquisition. In this paper, a Two-Piece-Ball (TPB) phantom is used to estimate a set of parameters that describe the geometry of a cone-beam CT system. Only multiple projections of the TPB phantom at one position are required, which can avoid the rotation errors when acquiring multi-angle projections. Also, a corresponding algorithm is derived. The performance of the method is evaluated through simulation and experimental data. The results demonstrated that the proposed method is valid and easy to implement. Furthermore, the experimental results from the Micro-CT system demonstrate the ability to reduce artifacts and improve image quality through geometric parameter calibration.

Wistar rats are widely used in biomedical research and commonly serve as a model organism in neuroscience studies. In most cases when noninvasive imaging is not used, studies assume a consistent baseline condition in rats that lack visible differences. While performing a series of traumatic brain injury studies, we discovered mild spontaneous ventriculomegaly in 70 (43.2%) of 162 Wistar rats that had been obtained from 2 different vendors. Advanced magnetic resonance (MR) imaging techniques, including MR angiography and diffusion tensor imaging, were used to evaluate the rats. Multiple neuropathologic abnormalities, including presumed arteriovenous malformations, aneurysms, cysts, white matter lesions, and astrogliosis were found in association with ventriculomegaly. Postmortem microcomputed tomography and immunohistochemical staining confirmed the presence of aneurysms and arteriovenous malformations. Diffusion tensor imaging showed significant decreases in fractional anisotropy and increases in mean diffusivity, axial diffusivity, and radial diffusivity in multiple white matter tracts (p < 0.05). These results could impact the interpretation, for example, of a pseudo-increase of axon integrity and a pseudo-decrease of myelin integrity, based on characteristics intrinsic to rats with ventriculomegaly. We suggest the use of baseline imaging to prevent the inadvertent introduction of a high degree of variability in preclinical studies of neurologic disease or injury in Wistar rats.

Embryogenesis is especially sensitive to external factors. The changes in its course are often used as biomarkers of environmental impact. Since spider embryogenesis takes place inside cocoons, it is crucial to find a reliable tool to analyze this developmental phase with no intrusion into the cocoons. The aim of this study was to verify the efficacy of X-ray microtomography for non-invasive analysis of embryonic morphology and egg quantity in the cocoons of Xerolycosa nemoralis and Agelena labyrinthica from polluted and reference sites. X-ray microtomography slice images as well as 3D images and animations obtained from digital visualization of those slides were used to study the morphology of embryos and egg arrangement in the cocoons. Any disorders in embryogenesis or malformation of embryos in relation to site of origin have not been found, but inside an egg cocoon of X. nemoralis from the polluted site embryos differing form each other by one developmental stage were identified. Egg calculation revealed a K- reproductive strategy of X. nemoralis from polluted sites. Finally, future prospects and benefits, and weaknessess of this method for the study of spider cocoons have presented., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Introduction: The purpose of this study was to investigate the ability of swept-source optical coherence tomographic (SS-OCT) imaging to detect a second mesiobuccal canal (MB2) in maxillary molars compared with visual inspection (VI) and dental operating microscopy (DOM)., Methods: Forty extracted human maxillary molars were examined. After the removal of half the crown and access cavity preparation, the existence of MB2 canals was confirmed based on cross-sectional images of each tooth obtained by micro-computed tomographic scanning as the reference standard. Then, the pulp chamber floor was scanned by SS-OCT imaging. Three dentists independently evaluated the SS-OCT images and the pulp chamber floor under VI and DOM for the presence of MB2 canals. The detection rate of MB2 canals for SS-OCT imaging, VI, and DOM was calculated, and statistical analysis was performed., Results: MB2 canals existed in 19 of 40 teeth (47.5%) using micro-CT imaging. Sensitivity of DOM (0.947) was significantly higher than that of SS-OCT imaging (0.632). Specificity of SS-OCT imaging (0.714) was significantly higher than that of DOM (0.333). No statistically significant differences were found for accuracy among the 3 methods. Kappa values of SS-OCT, VI, and DOM were 0.526, 0.417, and 0.326, respectively., Conclusions: SS-OCT imaging is noninvasive, involves no ionizing radiation, and is accurate for the detection of MB2 canals., (Copyright © 2014 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.)

Correlated imaging is the process of imaging a specimen with two complementary modalities and then registering and overlaying the fields obtained in each modality to create a composite view. One of the images is made somewhat transparent, allowing detail in the underlying image to be visible and assisting in the registration of the two images. As an example, an image localizing a specific tissue component by fluorescence may be overlaid atop a TEM image of the same field. The resulting composite image would demonstrate specific ultrastructural features in the high-resolution TEM field, which are colorized in the overlay. Other examples include composites from MicroCT or soft X-ray images overlaid atop light microscopy or TEM images. Automated image registration may be facilitated by a variety of sophisticated computer programs utilized by high-throughput laboratories. This chapter is meant for the more occasional user wishing to align images manually. ImageJ is a public domain, image processing program developed at the National Institutes of Health and is available to anyone as a free download. ImageJ performs marvelously well for the purpose of image registration; therefore, step-by-step instructions are included here. Specimen handling, including fixation and choice of embedding media, is not straightforward for correlative imaging. A step-by-step description of the protocols which work in our laboratory is included for simultaneous localization in LM, EM and micro-CT, as well as maintaining GFP emission in tissue embedded for TEM., (© 2014 Elsevier Inc. All rights reserved.)

Subject motion during high-resolution peripheral quantitative computed tomography (HR-pQCT) causes image artifacts that affect morphological analysis of bone quality. The aim of our study was to determine effectiveness of techniques for quality control in the presence of motion in vivo including automated and manual approaches. First, repeatability of manual grading was determined within and between laboratories. Given proper training using a standardized scale and training images (provided by the manufacturer), we found that manual grading is suitable for repeatable image quality grading within and across sites (ICC>0.7). Both a new automated technique providing motion measures based on projection moments, and traditional manual grading (1=best, 5=worst) were subsequently used to assess subject data for motion in N=137 image pairs (scan/re-scan) from the Canadian Multicentre Osteoporosis Study (CaMos) Calgary cohort. High quality image pairs were selected and measurement precision was estimated by calculating the coefficient of variation (CV). Consistent with previous data, density parameters (e.g. total bone mineral density) are more robust than structural (e.g. trabecular number) or finite element parameters (e.g. failure load). To obtain acceptable measurement precision, images should not exceed a manual grading of 3 (on a scale from 1 to 5) or an automatic (ε(T)) grading of 1.2. Automatic and manual grading provide comparable quality control, but the advantage of the automated technique is its ability to provide a motion value at scan time (providing a basis for real time decision regarding re-scan requirements), and the assessment is objective. Notably, automatic motion measurement can be performed retrospectively based on original scan data, and is therefore well suited for multi-center studies as well as any research where objective quality control is paramount., (Copyright © 2012 Elsevier Inc. All rights reserved.)

In order to improve realism in middle ear (ME) finite-element modeling (FEM), comprehensive and precise morphological data are needed. To date, micro-scale X-ray computed tomography (μCT) recordings have been used as geometric input data for FEM models of the ME ossicles. Previously, attempts were made to obtain these data on ME soft tissue structures as well. However, due to low X-ray absorption of soft tissue, quality of these images is limited. Another popular approach is using histological sections as data for 3D models, delivering high in-plane resolution for the sections, but the technique is destructive in nature and registration of the sections is difficult. We combine data from high-resolution μCT recordings with data from high-resolution orthogonal-plane fluorescence optical-sectioning microscopy (OPFOS), both obtained on the same gerbil specimen. State-of-the-art μCT delivers high-resolution data on the 3D shape of ossicles and other ME bony structures, while the OPFOS setup generates data of unprecedented quality both on bone and soft tissue ME structures. Each of these techniques is tomographic and non-destructive and delivers sets of automatically aligned virtual sections. The datasets coming from different techniques need to be registered with respect to each other. By combining both datasets, we obtain a complete high-resolution morphological model of all functional components in the gerbil ME. The resulting 3D model can be readily imported in FEM software and is made freely available to the research community. In this paper, we discuss the methods used, present the resulting merged model, and discuss the morphological properties of the soft tissue structures, such as muscles and ligaments.

Objectives: to evaluate the accuracy of cone beam computed tomography (CBCT) for volumetric measurement of teeth, using micro-computed tomography (Micro-CT) as the reference standard., Setting and Sample Population: The Department of Orthodontics at Sichuan University. The sample consisted of 27 maxillary and mandibular premolars of 15 patients, planned to be extracted for orthodontic treatment., Material and Methods: The 27 teeth were subjected to standardized CBCT scanning before extraction and Micro-CT scanning after extraction. From CBCT data, teeth were tissue segmented and then three-dimensionally (3D) reconstructed, while from Micro-CT data, teeth were 3D reconstructed directly. Tooth volumes were then calculated. The intra-observer repeatability and reproducibility of two observers and the overall between-instrument agreement of the measurements were evaluated using intra-class correlation coefficients (ICCs) and concordance correlation coefficient (CCC), respectively., Results: The intra-observer repeatability was high for both observers. ICCs were 0.999 and 0.998, respectively. The reproducibility of the two observers was also high (ICC, 0.740). The overall between-instrument agreement of the measurements was good, and CCC was 0.993 and its lower 95% confidence interval was 0.989., Conclusions: The accuracy of the CBCT method for volumetric measurement of teeth in vivo is comparable to the Micro-CT method in vitro. The CBCT method has the potential possibility to be applied in studies on root resorption associated with orthodontic force. Further study is needed to prove the sensitivity of the method., (© 2011 John Wiley & Sons A/S.)

In vivo microCT arthrography (µCTa) can be used to measure both quantity (volumetric) and quality (glycosaminoglycan content) of cartilage. This study investigated the accuracy of four segmentation techniques to isolate cartilage from µCTa datasets and then used the most accurate one to investigate if the µCTa method could show osteoarthritic changes in rat models during longitudinal follow-up. Volumetric measurements and glycosaminoglycan contents of patellar cartilage from in vivo µCTa-scans were compared with an ex vivo gold standard µCT-scan. Cartilage was segmented with three global thresholds and one local threshold algorithm. Comparisons were made for healthy and osteoarthritic cartilage. Next, three rat models were investigated for 24 weeks using µCTa. Osteoarthritis was induced by injection with a chemical (mono-iodoacetate), a surgical intervention (grooves applied in articular cartilage), and via exercise (strenuous running). After euthanasia, all knee joints were isolated for histology. Local thresholds accurately segmented cartilage from in vivo µCTa scans and best measured cartilage quantity and glycosaminoglycan content. Each of the three osteoarthritic rat models showed a specific pattern of osteoarthritis progression. All µCTa results were comparable to histology. In vivo µCTa is a sensitive technique for imaging cartilage degradation. Local thresholds enhanced the sensitivity of this method and will probably more accurately detect disease-modulating effects from interventional strategies. The data from rat models may serve as a reference for the time sequence of cartilage degeneration during in vivo testing of new strategies in osteoarthritis treatment., (Copyright © 2011 Orthopaedic Research Society.)

The agreement between measurements and the relative performance reproducibility among different microcomputed tomography (microCT) systems, especially at voxel sizes close to the limit of the instruments, is not known. To compare this reproducibility 3D morphometric analyses of mouse cancellous bone from distal femoral epiphyses were performed using three different ex vivo microCT systems: GE eXplore Locus SP, Scanco μCT35 and Skyscan 1172. Scans were completed in triplicate at 12 μm and 8 μm voxel sizes and morphometry measurements, from which relative values and dependence on voxel size were examined. Global and individual visually assessed thresholds were compared. Variability from repeated scans at 12 μm voxel size was also examined. Bone volume fraction and trabecular separation values were similar, while values for relative bone surface, trabecular thickness and number varied significantly across the three systems. The greatest differences were measured in trabecular thickness (up to 236%) and number (up to 218%). The relative dependence of measurements on voxel size was highly variable for the trabecular number (from 0% to 20% relative difference between measurements from 12 μm and 8 μm voxel size scans, depending on the system). The intra-system reproducibility of all trabecular measurements was also highly variable across the systems and improved for BV/TV in all the systems when a smaller voxel size was used. It improved using a smaller voxel size in all the other parameters examined for the Scanco system, but not consistently so for the GE or the Skyscan system. Our results indicate trabecular morphometry measurements should not be directly compared across microCT systems. In addition, the conditions, including voxel size, for trabecular morphometry studies in mouse bone should be chosen based on the specific microCT system and the measurements of main interest., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Micro-CT with a high spatial resolution in combination with computer-based-reconstruction techniques is considered a powerful tool for morphological study of insects. The quality of CT images crucially depends on the precise knowledge of the scan geometry of the micro-CT system. In this paper, we have proposed a method to calculate the deviation of rotating axis for compensating deficiency of existing methods. A practical application of this geometric calibration method of the micro-CT system for insect imaging is presented. We have performed the computer-simulation study and experimental study with our prototype micro-CT system. The results demonstrate that the proposed technique is accurate and robust. In addition, we have evaluated the imaging characteristics of the detector in terms of modulation-transfer function (MTF). Finally, insect imaging performance and image reconstruction from data acquired with different energies are presented.

Synchrotron-radiation-based microcomputed-tomography (SR-μCT) is a powerful tool for yielding 3D structural information of high spatial and contrast resolution about a specimen preserved in its natural state. A large number of projection views are required currently for yielding SR-μCT images by use of existing algorithms without significant artifacts. When a wet biological specimen is imaged, synchrotron x-ray radiation from a large number of projection views can result in significant structural deformation within the specimen. A possible approach to reducing imaging time and specimen deformation is to decrease the number of projection views. In the work, using reconstruction algorithms developed recently for medical computed tomography (CT), we investigate and demonstrate image reconstruction from sparse-view data acquired in SR-μCT. Numerical results of our study suggest that images of practical value can be obtained from data acquired at a number of projection views significantly lower than those used currently in a typical SR-μCT imaging experiment., (© 2011 American Institute of Physics)

Use of high-resolution micro-computed tomography (microCT) imaging to assess trabecular and cortical bone morphology has grown immensely. There are several commercially available microCT systems, each with different approaches to image acquisition, evaluation, and reporting of outcomes. This lack of consistency makes it difficult to interpret reported results and to compare findings across different studies. This article addresses this critical need for standardized terminology and consistent reporting of parameters related to image acquisition and analysis, and key outcome assessments, particularly with respect to ex vivo analysis of rodent specimens. Thus the guidelines herein provide recommendations regarding (1) standardized terminology and units, (2) information to be included in describing the methods for a given experiment, and (3) a minimal set of outcome variables that should be reported. Whereas the specific research objective will determine the experimental design, these guidelines are intended to ensure accurate and consistent reporting of microCT-derived bone morphometry and density measurements. In particular, the methods section for papers that present microCT-based outcomes must include details of the following scan aspects: (1) image acquisition, including the scanning medium, X-ray tube potential, and voxel size, as well as clear descriptions of the size and location of the volume of interest and the method used to delineate trabecular and cortical bone regions, and (2) image processing, including the algorithms used for image filtration and the approach used for image segmentation. Morphometric analyses should be based on 3D algorithms that do not rely on assumptions about the underlying structure whenever possible. When reporting microCT results, the minimal set of variables that should be used to describe trabecular bone morphometry includes bone volume fraction and trabecular number, thickness, and separation. The minimal set of variables that should be used to describe cortical bone morphometry includes total cross-sectional area, cortical bone area, cortical bone area fraction, and cortical thickness. Other variables also may be appropriate depending on the research question and technical quality of the scan. Standard nomenclature, outlined in this article, should be followed for reporting of results., (2010 American Society for Bone and Mineral Research.)

Objectives: Radiological evaluation of the position of cochlear implant (CI) devices is an upcoming method for quality control after CI surgery. First, results of imaging of the middle and inner ear with digital volume tomography (DVT) show considerable advantages such as exceptional image quality, thin slice thickness, and low radiation dose. The aim of this study was to evaluate whether DVT is an appropriate method for postoperative imaging of CI patients and to identify the exact position of the implant array within the cochlear by multiple measurements., Design: Thirteen formalin-fixed temporal bone specimens were implanted with a CI array and scanned in DVT. To determine the exact electrode position, these specimens were ground and stained for microscopic measurements. The measurements on grindings acted as a referee and were compared with the measurements in DVT scans. The statistical analysis between the two measurement protocols was performed using the Bland-Altman method., Results: Best achievable agreement between DVT scans and histological reference was shown. Mean differences between DVT and grindings from -1.55 to -65.40 microm were calculated. All means are within the region of accuracy. General positioning of the implant into the cochlea could be verified in all specimens. The exact position of the implanted array within the cochlear scalae could be recognized correctly in 11 of 13 cases in DVT. It was possible to identify shiftings between the tympanic and vestibular scalae in all cases., Conclusion: DVT seems to be a convenient technique for postoperative position control after cochlear implantation.

We report a precise, simple, and accessible method of mathematically measuring and modeling the three-dimensional (3D) geometry of semicircular canals (SCCs) in living humans. Knowledge of this geometry helps understand the development and physiology of SCC stimulation. We developed a framework of robust techniques that automatically and accurately reconstruct SCC geometry from computed tomography (CT) images and are directly validated using micro-CT as ground truth. This framework measures the 3D centroid paths of the bony SCCs allowing direct comparison and analysis between ears within and between subjects. An average set of SCC morphology is calculated from 34 human ears, within which other geometrical attributes such as nonplanarity, radius of curvature, and inter-SCC angle are examined, with a focus on physiological implications. These measurements have also been used to critically evaluate plane fitting techniques that reconcile many of the discrepancies in current SCC plane studies. Finally, we mathematically model SCC geometry using Fourier series equations. This work has the potential to reinterpret physiology and pathophysiology in terms of real individual 3D morphology.

The aim of this work was to present and discuss a quality control protocol for in vitro micro-computed tomography (microCT), based on the adaptation of the quality control protocols for medical computed tomography. The importance of establishing a quality control protocol is related to the opportunity to identify problems on time comparing the microCT images acquired in different time points, and in this way to verify the performance of the device. The proposed quality control protocol was applied for a long-time monitoring period to verify the stability of the micro-tomographic system over time. The protocol proposed in this study was applied to the histomorphometric characterization of bone tissue, but it can be used on a wide range of in vitro microCT applications. Noise and uniformity tests, taken and adapted to micro-tomographic system by medical standard guidelines of quality control, were performed by the use of a water phantom. An accuracy test was designed and performed by the use of a morphometric calibrated phantom. All these tests were performed during a long-time monitoring period to control the stability of the system. Specific control charts and monitoring parameters for each test were used to represent the monthly measures collected during 20 months and an out of control condition was defined. The reference values (baseline), calculated to control the stability of micro-tomographic system over time, were calculated during acceptance/status test. During the period, no out of control conditions in noise, uniformity and accuracy tests were recorded. However, a changing condition was found in noise test, as showed by using statistical C (P < 0.01) and Kruskal-Wallis (P < 0.05) tests. In particular, a Wilcoxon rank sum test with Bonferroni correction (P < 0.0125) was applied in noise test to investigate which of the comparisons among first five acquisitions of year 2004 (group B.L.) and each group was significant (P < 0.0125). The noise showed a slight but significant increase over the years compared to baseline value; however, no out of control conditions were recorded. Nonetheless, a maintenance service to control the performance of mechanical components of microCT was required and performed.

Objective: To define the nature of structural bone changes in patients with rheumatoid arthritis (RA) compared with those in healthy individuals by using the novel technique of high-resolution microfocal computed tomography (micro-CT)., Methods: Fifty-eight RA patients and 30 healthy individuals underwent a micro-CT scan of the proximal wrist and metacarpophalangeal joints. Bone lesions such as cortical breaks, osteophytes, and surface changes were quantified on 2-dimensional (2-D) slices as well as by using 3-D reconstruction images, and exact localization of lesions was recorded., Results: Micro-CT scans could detect bone lesions <0.5 mm in width or depth. Small erosions could be observed in healthy individuals and RA patients, whereas lesions >1.9 mm in diameter were highly specific for RA. Cortical breaks were mostly found along the radial sites of the metacarpal heads. No significant difference in the presence of osteophytes between healthy individuals and RA patients was found. Cortical surface changes, presumably cortical thinning and fenestration, became evident from 3-D reconstructions and were more pronounced in RA patients., Conclusion: Micro-CT allows exact detection of morphologic changes of juxtaarticular bone in healthy individuals and RA patients. Even healthy individuals occasionally show bone changes, but the severity of these lesions, with the exception of osteophytes, is greater in RA patients. Thus, micro-CT allows accurate differentiation among physiologic bone changes in joints and among types of pathologic bone damage resulting from RA.

Background and Purpose: Animal models developed in rats and mice have become indispensable in preclinical cerebrovascular research. Points of interest include the investigation of the vascular bed and the morphology and function of the arterial, capillary, and venous vessels. Because of their extremely small caliber, in vivo examination of these vessels is extremely difficult. In the present study we have developed a method to provide fast 3D in vivo analysis of cerebral murine vessels using volume computed tomography-angiography (vCTA)., Methods: Using an industrial X-ray inspection system equipped with a multifocus cone beam X-ray source and a 12-bit direct digital flatbed detector, high-speed vCTA (180 degrees rotation in 40 s. at 30 fps) was performed in anesthetized mice. During the scan an iodinated contrast agent was infused via a tail vein. Images were reconstructed using a filtered backprojection algorithm. Image analysis was performed by maximum intensity projection (MIP) and 3D volume reconstruction., Results: All mice tolerated i.v. injection of the iodinated contrast agent well. Smallest achievable voxel size of raw data while scanning the whole neurocranium was 16 mum. Anatomy of cerebral vessels was assessable in all animals, and anatomic differences between mouse strains could easily be detected. Mean vessel diameter was measured in C57BL/6 and BALBc mice. Changes of vessel caliber were assessable by repeated vCTA., Conclusions: Ultra fast in vivo vCTA of murine cerebral vasculature is feasible at resolutions down to 16 mum. The technique allows the assessment of vessel caliber changes in living mice, thus providing an interesting tool to monitor different features such as vasospasm or vessel patency.