scholarly journals Three-Dimensional X-ray Imaging of β-Galactosidase Reporter Activity by Micro-CT: Implication for Quantitative Analysis of Gene Expression

2021 ◽  
Vol 11 (6) ◽  
pp. 746
Author(s):  
Olga Ermakova ◽  
Tiziana Orsini ◽  
Paolo Fruscoloni ◽  
Francesco Chiani ◽  
Alessia Gambadoro ◽  
...  
Keyword(s):  
Gene Expression ◽  
Quantitative Analysis ◽  
Three Dimensional ◽  
Micro Ct ◽  
Lacz Reporter ◽  
Lacz Reporter Gene ◽  
X Ray ◽  
Reporter Activity ◽  
X Ray Imaging ◽  
Murine Brain

Acquisition of detailed anatomical and molecular knowledge from intact biological samples while preserving their native three-dimensional structure is still a challenging issue for imaging studies aiming to unravel a system’s functions. Three-dimensional micro-CT X-ray imaging with a high spatial resolution in minimally perturbed naive non-transparent samples has recently gained increased popularity and broad application in biomedical research. Here, we describe a novel X-ray-based methodology for analysis of β-galactosidase (lacZ) reporter-driven gene expression in an intact murine brain ex vivo by micro-CT. The method relies on detection of bromine molecules in the product of the enzymatic β-galactosidase reaction. Enhancement of the X-ray signal is observed specifically in the regions of the murine brain where expression of the lacZ reporter gene is also detected histologically. We performed quantitative analysis of the expression levels of lacZ reporter activity by relative radiodensity estimation of the β-galactosidase/X-gal precipitate in situ. To demonstrate the feasibility of the method, we performed expression analysis of the Tsen54-lacZ reporter gene in the murine brain in a semi-quantitative manner. Human mutations in the Tsen54 gene cause pontocerebellar hypoplasia (PCH), a group of severe neurodegenerative disorders with both mental and motor deficits. Comparing relative levels of Tsen54 gene expression, we demonstrate that the highest Tsen54 expression is observed in anatomical brain substructures important for the normal motor and memory functions in mice.

scholarly journals X-ray 3D imaging of gene expression in whole-mount murine brain by microCT, implication for functional analysis of tRNA endonuclease 54 gene mutated in pontocerebellar hypoplasia

2019 ◽  
Author(s):  
Olga Ermakova ◽  
Tiziana Orsini ◽  
Paolo Fruscoloni ◽  
Francesco Chiani ◽  
Alessia Gambadoro ◽  
...  
Keyword(s):  
Gene Expression ◽  
Expression Pattern ◽  
Reporter Gene ◽  
Three Dimensional ◽  
Gene Expression Pattern ◽  
Lacz Gene ◽  
Pontocerebellar Hypoplasia ◽  
X Ray ◽  
Whole Mount ◽  
Murine Brain

AbstractAcquisition of detailed structural and molecular information from intact biological samples, while preserving cellular three-dimensional structures, still represents a challenge for biological studies aiming to unravel system functions. Here we describe a novel X-ray-based methodology for analysis of gene expression pattern in intact murine brain ex vivo by microCT. The method relays on detection of bromine molecules in the products of enzymatic reaction generated by the β-galactosidase (lacZ) gene reporter. To demonstrate the feasibility of the method, the analysis of the expression pattern of tRNA endonuclease 54 (Tsen54)-lacZ reporter gene in the whole-mount murine brain in semi-quantitative manner is performed. Mutations in Tsen54 gene causes pontocerebellar hypoplasia (PCH), severe neurodegenerative disorder with both mental and motor deficits. Comparing relative levels of Tsen54 gene expression, we have demonstrated that highest Tsen54 expression observed in anatomical brain substructures important for the normal motor and memory functions in mice. In the forebrain strong expression in perirhinal, retrosplenial and secondary motor areas was observed. In olfactory area Tsen54 is highly expressed in the nucleus of the lateral olfactory tract, anterior olfactory and bed nuclei, while in hypothalamus in lateral mammillary nucleus and preoptic area. In hindbrain Tsen54 is expressed in the reticular, cuneate and trigeminal nuclei of medulla, and in pontine gray of pons and in cerebellum, in the molecular and Purkinje cell layers. Delineating anatomical brain regions in which Tsen54 is strongly expressed will allow functionally address the role Tsen54 gene in normal physiology and in PCH disease.Significance StatementCharacterization of gene expression pattern in the brain of model organisms is critical for unravelling the gene function in normal physiology and disease. It is performed by optical imaging of the two-dimensional brain sections which then assembled in volume images. Here we applied microCT platform, which allows three-dimensional imaging of non transparent samples, for analysis of gene expression. This method based on detection by X-ray the bromine molecules presented in the products generated by enzymatic activity of b-galactosidase reporter gene. With this method we identify anatomical brain substructures in which Tsen54 gene, mutated in pontocerebellar hypoplasia disease, is expressed.

X-ray imaging with ultra-small-angle X-ray scattering as a contrast mechanism

2004 ◽  
Vol 37 (5) ◽  
pp. 757-765 ◽  
Author(s):  
L. E. Levine ◽  
G. G. Long
Keyword(s):  
Small Angle ◽  
Imaging Technique ◽  
Three Dimensional ◽  
Sample Volume ◽  
Contrast Imaging ◽  
X Ray ◽  
X Ray Scattering ◽  
X Ray Imaging ◽  
Contrast Mechanism ◽  
Ray Scattering

A new transmission X-ray imaging technique using ultra-small-angle X-ray scattering (USAXS) as a contrast mechanism is described. USAXS imaging can sometimes provide contrast in cases where radiography and phase-contrast imaging are unsuccessful. Images produced at different scattering vectors highlight different microstructural features within the same sample volume. When used in conjunction with USAXS scans, USAXS imaging provides substantial quantitative and qualitative three-dimensional information on the sizes, shapes and spatial arrangements of the scattering objects. The imaging technique is demonstrated on metal and biological samples.

Interfacial Structure Control and Three-Dimensional X-ray Imaging of an Epoxy Monolith Bonding System with Surface Modification

Langmuir ◽  
2020 ◽  
Vol 36 (37) ◽  
pp. 10923-10932
Author(s):  
Nanako Sakata ◽  
Yoshihiro Takeda ◽  
Masaru Kotera ◽  
Yasuhito Suzuki ◽  
Akikazu Matsumoto
Keyword(s):  
Surface Modification ◽  
Three Dimensional ◽  
Interfacial Structure ◽  
X Ray ◽  
Structure Control ◽  
Bonding System ◽  
X Ray Imaging

Three Dimensional X-Ray Computed Tomography in Materials Science

MRS Bulletin ◽  
1988 ◽  
Vol 13 (1) ◽  
pp. 13-18 ◽  
Author(s):  
J.H. Kinney ◽  
Q.C. Johnson ◽  
U. Bonse ◽  
M.C. Nichols ◽  
R.A. Saroyan ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
High Resolution ◽  
Sample Preparation ◽  
Visible Light ◽  
Three Dimensional ◽  
Materials Characterization ◽  
Imaging Technology ◽  
X Ray ◽  
X Ray Imaging ◽  
Visible Light Imaging

Imaging is the cornerstone of materials characterization. Until the middle of the present century, visible light imaging provided much of the information about materials. Though visible light imaging still plays an extremely important role in characterization, relatively low spatial resolution and lack of chemical sensitivity and specificity limit its usefulness.The discovery of x-rays and electrons led to a major advance in imaging technology. X-ray diffraction and electron microscopy allowed us to characterize the atomic structure of materials. Many materials vital to our high technology economy and defense owe their existence to the understanding of materials structure brought about with these high-resolution methods.Electron microscopy is an essential tool for materials characterization. Unfortunately, electron imaging is always destructive due to the sample preparation that must be done prior to imaging. Furthermore, electron microscopy only provides information about the surface of a sample. Three dimensional information, of great interest in characterizing many new materials, can be obtained only by time consuming sectioning of an object.The development of intense synchrotron light sources in addition to the improvements in solid state imaging technology is revolutionizing materials characterization. High resolution x-ray imaging is a potentially valuable tool for materials characterization. The large depth of x-ray penetration, as well as the sensitivity of absorption crosssections to atomic chemistry, allows x-ray imaging to characterize the chemistry of internal structures in macroscopic objects with little sample preparation. X-ray imaging complements other imaging modalities, such as electron microscopy, in that it can be performed nondestructively on metals and insulators alike.

High-Speed X-Ray Imaging of Diesel Injector Needle Motion

2009 ◽  
Author(s):  
A. L. Kastengren ◽  
C. F. Powell ◽  
Z. Liu ◽  
K. Fezzaa ◽  
J. Wang
Keyword(s):  
High Speed ◽  
Three Dimensional ◽  
Linear Increase ◽  
Nozzle Geometry ◽  
X Ray ◽  
Needle Position ◽  
X Ray Imaging ◽  
Linear Oscillation ◽  
Degree Of Convergence

Phase-enhanced x-ray imaging has been used to examine the geometry and dynamics of four diesel injector nozzles. The technique uses a high-speed camera, which allows the dynamics of individual injection events to be observed in real time and compared. Moreover, data has been obtained for the nozzles from two different viewing angles, allowing for the full three-dimensional motions of the needle to be examined. This technique allows the needle motion to be determined in situ at the needle seat and requires no modifications to the injector hardware, unlike conventional techniques. Measurements of the nozzle geometry have allowed the average nozzle diameter, degree of convergence or divergence, and the degree of rounding at the nozzle inlet to be examined. Measurements of the needle lift have shown that the lift behavior of all four nozzles consists of a linear increase in needle lift with respect to time until the needle reaches full lift and a linear decrease as the needle closes. For all four nozzles, the needle position oscillates at full lift with a period of 170–180 μs. The full-lift position of the needle changes as the rail pressure increases, perhaps reflecting compression of the injector components. Significant lateral motions were seen in the two single-hole nozzles, with the needle motion perpendicular to the injector axis resembling a circular motion for one nozzle and linear oscillation for the other nozzle. The two VCO multihole nozzles show much less lateral motion, with no strong oscillations visible.

Parallelized Bayesian inversion for three-dimensional dental X-ray imaging

2006 ◽  
Vol 25 (2) ◽  
pp. 218-228 ◽  
Author(s):  
V. Kolehmainen ◽  
A. Vanne ◽  
S. Siltanen ◽  
S. Jarvenpaa ◽  
J.P. Kaipio ◽  
...  
Keyword(s):  
Three Dimensional ◽  
Bayesian Inversion ◽  
X Ray ◽  
X Ray Imaging

Three-dimensional x-ray imaging with off-axis x-ray zone plate

1995 ◽  
Author(s):  
Heung-Rae Lee ◽  
E. Anderson ◽  
L. B. Da Silva ◽  
J. E. Trebes
Keyword(s):  
Three Dimensional ◽  
Zone Plate ◽  
X Ray ◽  
X Ray Imaging

scholarly journals Micro-CT screening of old shell collections helps to understand the distribution of viviparity in the highly diversified clausiliid clade of land snails

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Anna Sulikowska-Drozd ◽  
Piotr Duda ◽  
Katarzyna Janiszewska
Keyword(s):  
Nineteenth Century ◽  
Reproductive Strategy ◽  
Southeast Asian ◽  
Land Snails ◽  
Micro Ct ◽  
X Ray ◽  
Ct Screening ◽  
Reproductive Modes ◽  
X Ray Imaging

AbstractCurrent zoological research may benefit in many ways from the study of old collections of shells. These collections may provide materials for the verification of broad zoogeographical and ecological hypotheses on the reproduction of molluscs, as they include records from many areas where sampling is currently impossible or very difficult due to political circumstances. In the present paper we present data on viviparous and embryo-retention reproductive modes in clausiliid land snails (subfamily Phaedusinae) acquired from specimens collected since the nineteenth century in the Pontic, Hyrcanian, and East and Southeast Asian regions. X-ray imaging (micro-CT) enabled relatively quick screening of more than 1,000 individuals classified within 141 taxa, among which we discovered 205 shells containing embryos or eggs. Gravid individuals were found to belong to 55 species, representing, for some of these species, the first indication of brooding reproductive strategy.

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