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Differential diagnosis of intracerebral hemorrhage due to traumatic and non-traumatic origin is a challenging issue, especially in the absence of visible body injuries and other signs of violence. For a forensic expert-practitioner, the main thing is objectivity, accuracy, and speed of obtaining the result, which could fully satisfy the methods of laser polarimetry in the case of differential diagnosis of hemorrhages of traumatic and non-traumatic origin in the human brain matter. The purpose: To develop forensic criteria for the differential diagnosis of intracerebral hemorrhage of traumatic and non-traumatic origin using Mueller-matrix microscopy of linear dichroism. Materials and methods: The object of the study was brain matter samples, collected in 115 corpses of both sexes aged 22 to 86 years with accurately known causes of death. The causes of death included coronary artery disease, traumatic brain hemorrhage, ischemic brain infarction, and non-traumatic brain hemorrhages. Brain matter samples have been frozen rapidly, and histological sections have been made using a freezing microtome. The slices have been evaluated using the method of azimuthal-invariant Mueller-matrix microscopy, followed by mathematical and statistical processing of the results. Results: significant topographic heterogeneity of Mueller-matrix invariants of brain matter samples linear dichroism maps among all research groups have been found. Individual and significant variations of dispersion, asymmetry, and kurtosis magnitude have confirmed the heterogeneity of Mueller-matrix invariants. The difference of images and histograms among groups is explained by the presence of necrotic changes in the case of ischemic brain necrosis and blood cells in the brain matter samples. For the set of central statistical moments of the 1st, 3rd, and 4th orders, which characterize the distributions of circular dichroism, the method of MM-microscopy in the differentiation of samples of non-hemorrhage and hemorrhage groups reaches a satisfactory level - 78% - 84%. Conclusions: Brain matter samples Muller-matrix microscopy of linear dichroism is effective for solving diagnostic problems of forensic medicine related to evaluating the cause of death from intracerebral hemorrhage of various origins. Diagnostic efficiency of Mueller-matrix mapping of polarization manifestations of linear dichroism method for intergroup differentiation of samples of deaths from traumatic hemorrhage (Group 2) and ischemic cerebral infarction (Group 3) reaches a satisfactory level of 79 - 84%.

PURPOSE Precise correlation between three-dimensional (3D) imaging and histology can aid biomechanical modeling of the breast. We develop a framework to register ex vivo images to histology using a novel cryo-fluorescence tomography (CFT) device. METHODS A formalin-fixed cadaveric breast specimen, including chest wall, was subjected to high-resolution magnetic resonance (MR) imaging. The specimen was then frozen and embedded in an optimal cutting temperature (OCT) compound. The OCT block was placed in a CFT device with an overhead camera and 50 μm thick slices were successively shaved off the block. After each shaving, the block-face was photographed. At select locations including connective/adipose tissue, muscle, skin, and fibroglandular tissue, 20 μm sections were transferred onto cryogenic tape for manual hematoxylin and eosin staining, histological assessment, and image capture. A 3D white-light image was automatically reconstructed from the photographs by aligning fiducial markers embedded in the OCT block. The 3D MR image, 3D white-light image, and photomicrographs were rigidly registered. Target registration errors (TREs) were computed based on 10 pairs of points marked at fibroglandular intersections. The overall MR-histology registration was used to compare the MR intensities at tissue extraction sites with a one-way analysis of variance. RESULTS The MR image to CFT-captured white-light image registration achieved a mean TRE of 0.73 ± 0.25 mm (less than the 1 mm MR slice resolution). The block-face white-light image and block-face photomicrograph registration showed visually indistinguishable alignment of anatomical structures and tissue boundaries. The MR intensities at the four tissue sites identified from histology differed significantly (p

Background. The quantity and quality of the etching pattern produced by acids play a significant role in the wettability and contact angle of the adhesive with the enamel surface in orthodontics. The better the etch pattern, the more the surface energy of the enamel, better the penetration of the adhesive, which ultimately results in better bond strength. The present study aimed to evaluate the contact angle of the bonding agent with the enamel surface etched by five commercially available etchants and check if any difference existed between the five. Materials and Methods. Twenty-five human maxillary and mandibular central incisors and premolars extracted for orthodontic or dental purposes were used in this study as samples. The teeth were allocated into five groups based upon the etchant used to etch the enamel surface. After the samples were etched, a hard tissue microtome was used to create thin slices of the enamel surface. The samples were then exposed to bonding agent Ormco Enlight. The bonding agent was released in the form of droplets onto the enamel slices mounted on a contact angle goniometer. The contact angle values were tabulated, and statistical analysis using the one-way ANOVA test was carried out. Results. The contact angle measurements of the etchant group, DPI, were the lowest, while D-tech has the highest contact angle values. However, statistical analysis revealed no statistically significant difference between the contact angle measurements of the five groups included in the study. Conclusion. No statistically significant difference existed between the five etchant groups included in the study. However, commercially available etchant DPI showed lesser contact angle and thus better wettability in comparison to other groups. Further elemental analysis and surface analysis are required to validate these results.

Paraffin histology is one of the most important and commonly-used laboratory techniques in diagnostic histopathology. The discovery of paraffin embedding is often attributed to the pathologist Edwin Klebs. Klebs was following the lead of Stricker, who embedded embryos in a mixture of hot stearin and white beeswax. We show that Klebs experimented with paraffin wax for embedding tumour tissue. But he quickly rejected it as unsuitable because paraffin wax did not infiltrate the tissue. One of Klebs' correspondents, embryologist Wilhelm His, Sr., learned of Klebs' experiments and decided to try paraffin embedding. His dehydrated chicken embryos in alcohol, cleared them in lavender oil, and dripped hot paraffin wax onto them. This process allowed His to cut good sections. Here, we have replicated His's paraffin embedding protocol in order to determine whether His had indeed made the landmark discovery of infiltration embedding with paraffin wax. We followed the protocol that he gives in his 1868 monograph on the early development of the chicken. The protocol described by His failed, in our hands, to yield sections of the quality that he illustrates in his monograph. Typically, the tissue disintegrated when sectioned due to poor infiltration of the wax. Usable sections could only be obtained if His's protocol was modified by melting the embedded embryos in fresh paraffin wax. One explanation for our findings is that we failed to faithfully replicate His's protocol. Another is that his protocol was incomplete. We suggest that His is likely to have discovered and perfected infiltration embedding with paraffin wax but did not publish a complete protocol.Die paraffinbasierte histologische Untersuchung ist eines der wichtigsten angewendeten Laborverfahren in der diagnostischen Histopathologie. Die Entwicklung der Paraffineinbettung wird oft dem Pathologen Edwin Klebs zugeschrieben. Klebs orientierte sich an den Arbeiten von Stricker, der Embryonen in eine Mischung aus heißem Tristearin und weißem Bienenwachs einbettete, und experimentierte mit Paraffin, um Tumorgewebe einzubetten. Er verwarf den Ansatz aber rasch, da Paraffin das Gewebe nicht infiltrierte. Klebs stand im Briefwechsel mit dem Embryologen Wilhelm His Senior, der von dessen Experimenten erfuhr und beschloss, sich an der Paraffineinbettung zu versuchen. His entwässerte Hühnerembryonen in Alkohol, klärte sie mit Lavendelöl und träufelte heißes Paraffin darüber. Dieses Verfahren ermöglichte ihm die Anfertigung guter Schnitte. In der vorliegenden Arbeit haben wir His’ Protokoll der Paraffineinbettung nachvollzogen, um zu ermitteln, ob His tatsächlich die Infiltrationseinbettung mit Paraffin gelungen ist. Wir befolgten das Protokoll aus seiner 1868 erschienenen Monografie. Anhand der Anweisungen von His gelang es uns nicht, Schnitte mit der von ihm illustrierten Qualität herzustellen. In der Regel zerfiel das Gewebe beim Schneiden wegen schlechter Wachsinfiltration. Brauchbare Schnitte wurden nur bei Abwandlung von His’ Protokoll durch das Wiedereinschmelzen der eingebetteten Embryos in frischem Paraffin erreicht. Möglicherweise ist es uns nicht gelungen ist, das Protokoll von His detailgetreu zu replizieren, oder das Protokoll war unvollständig. Nach unserer Einschätzung hat His wahrscheinlich die Infiltrationseinbettung mit Paraffin erfunden und perfektioniert, aber kein vollständiges Protokoll veröffentlicht.

Objective: The aim of this study is to evaluate histologically the healing potential of injectable macro-porous Calcium Phosphate Cement and autogenous bone. (Normal bone healing) on surgically induced bone defect on femur bone of rabbit. Materials and Methods: Twelve adult white male New Zealand rabbits were selected for this study. Rabbits were divided into two groups and the two groups according to a two evaluation periods divided into 2 subgroups. Rabbits were anesthetized with intramuscular anethesia, and two osseous defects had been created in the distal aspect of the femur bone.one of this defects injected by a grafting material (SI-HPMC CPC) and the other left empty as control (normal healing).Animal were sacrificed after two sacrifice dates and the femur bone excised for histological and statistical evaluation.esults: Rabbits from each group were sacrificed after 2 and 4 weeks. The femur bone containing the induced defect was dissected. Each femur bone was excised using hard tissue microtome. Each slice was then fixed in 10% neutral buffered formalin. The formalin-fixed bone samples were decalcified in 15% buffered formic acid solution and processed for routine histological examination using hematoxylin and eosin stain under light microscope.. Conclusion: According to our results in the present study, we can consider Si-HPMC CPC a viable alternative to the autogenous bone in the healing of osseous bone defects . In the present study, the new and simple method to prepare macroporous CPCs using Si-HPMC a foaming agent in connected syringe , result

Aim: The aim of the study is to evaluate the inhibitory effect of single-bottle and two-bottle self-etch adhesive system after surface pretreatment with 6.5% proanthocyanidin (PA) on cysteine cathepsins (CCs) present in dentin hybrid layer using gelatin zymography. Materials and Methods: Thirty-five samples of dentin measuring 3 mm × 3 mm were obtained from 35 caries-free human third molars which were divided into seven groups. The bonded specimens were cut vertically into 1 mm thick adhesive/dentin interfaces with microtome and were pulverized into powder and subjected to zymography analysis for enzymatic activity. Results: Group VI and VII which were positive control groups had shown thicker clear bands of 50 kDa. Group IV and V which had the administration of chlorhexidine showed thicker clear bands of 50 kDa which indicated slight activity of CCs. Group II in which PA used along with single-bottle system also showed thicker clear band of 59 kDa. CCs could be active due to strong acidic nature of monomer in this group. Group III in which PA used along with two bottle system had no band formation where there is crosslinking of catalytic and allosteric domains of enzymes by PA. Conclusion: Under the limitations of this study, PA proved as a non-specific natural inhibitor of CCs. Two-bottle system showed inhibition of CCs and single bottle system failed to inhibit their activity. Future research have to be done on the bond strength and durability of self-etch adhesives after using PA and its effect on long term survival of the restorations.

Aim of the study: Using the example of a clinical case, to show the importance of dynamic observation by means of various diagnostic methods of intestinal imaging, in combination with a pathomorphological confirmation method for setting the correct clinical diagnosis. Materials and methods: The studies were performed on multislice computed tomographs Brilliance CT and ICT (Philips Medical Systems) and magnetic resonance imaging machines (GE), with intravenous contrast enhancement. We used special methods of contrasting the lumen of the intestine with gradual and retrograde filling it with water. The surgical material fixed in 10% neutral formuline for 10-24 hours. Tissue fragments were processed in a Leica TP1020 histoprocessor (Leica BioSystems, Germany) according to a standard protocol, after which it was embedded in paraffin blocks. Histological sections with a thickness of 3-5 μm were made by a Leica RM2245 microtome (Leica BioSystems, Germany). The prepared histological preparations were stained with hematoxylin and eosin in a Leica Autosteiner XL histostiner (Leica BioSystems, Germany). Results: From 2013 to 2020 patient K. underwent 4 colonoscopies, 5 esophagogastroduodenoscopies, 1 Х-ray examination with small bowel series, 3 MRI studies and 5 CT studies, including those with intravenous contrast enhancement, with a total radiation exposure of 187 mSv. The diagnosis was revised from ulcerative colitis, Crohn’s disease to NC. The final diagnosis was made by the pathologist’s analysis of materials. Conclusion: The clinical case presented by us clearly demonstrates the importance of dynamic observation by computed and magnetic resonance imaging, in combination with a pathomorphological confirmation method.

Abdominal sections of honeybees undergo numerous reciprocating motions during their lifetime. However, the overlapped contact areas adjacent to the abdominal sections have a shallow wear extent, a physical mechanism that remains obscure to date. Therefore, this study explored a biofrictional reduction model based on a solid surface texture and the hairy surface of the honeybee abdomen. We collected honeybee samples and observed their abdomens using a camera (Zeiss Stemi 508). Subsequently, we sliced these samples using a microtome and detected their microscopic friction. The exterior surface of the honeybee abdomen was not smooth but was distributed with a dense microvilli structure, which played a vital role in adjusting the friction reduction characteristics between the abdominal sections. When the adjacent abdominal sections moved relatively to each other, their upper and lower surfaces were not in direct rigid contact. Briefly, this study shows that the microscale hair arrays on the surface of the posterior abdominal segment can significantly reduce real contact area and friction, which considerably decreases wear or abrasion. The friction reduction mechanism alleviates the abrasion during the relative bending movement and saves a large amount of energy, which is essential for the honeybees' daily activities. This microtexture compliance friction reduction characteristic could be used to fabricate hierarchical surfaces for long-lasting friction reduction mechanisms, which increase the life of soft devices, including soft actuators and hinges.

Paraffin histology is one of the most important and commonly used laboratory techniques in diagnostic histopathology. The discovery of paraffin embedding is often attributed to the pathologist Edwin Klebs. Klebs was following the lead of Stricker, who embedded embryos in a mixture of hot stearin and white beeswax. We show that Klebs experimented with paraffin wax for embedding tumour tissue. But he quickly rejected it as unsuitable because paraffin wax did not infiltrate the tissue. One of Klebs' correspondents, embryologist Wilhelm His, Sr., learned of Klebs' experiments and decided to try paraffin embedding. His dehydrated chicken embryos in alcohol, cleared them in lavender oil, and dripped hot paraffin wax onto them. This process allowed His to cut good sections. Here, we have replicated His's paraffin embedding protocol in order to determine whether His had indeed made the landmark discovery of infiltration embedding with paraffin wax. We followed the protocol that he gives in his 1868 monograph on the early development of the chicken. The protocol described by His failed, in our hands, to yield sections of the quality that he illustrates in his monograph. Typically, the tissue disintegrated when sectioned due to poor infiltration of the wax. Usable sections could only be obtained if His's protocol was modified by melting the embedded embryos in fresh paraffin wax. One explanation for our findings is that we failed to faithfully replicate His's protocol. Another is that his protocol was incomplete. We suggest that His is likely to have discovered and perfected infiltration embedding with paraffin wax but did not publish a complete protocol.Die paraffinbasierte histologische Untersuchung ist eines der wichtigsten angewendeten Laborverfahren in der diagnostischen Histopathologie. Die Entwicklung der Paraffineinbettung wird oft dem Pathologen Edwin Klebs zugeschrieben. Klebs orientierte sich an den Arbeiten von Stricker, der Embryonen in eine Mischung aus heißem Tristearin und weißem Bienenwachs einbettete, und experimentierte mit Paraffin, um Tumorgewebe einzubetten. Er verwarf den Ansatz aber rasch, da Paraffin das Gewebe nicht infiltrierte. Klebs stand im Briefwechsel mit dem Embryologen Wilhelm His Senior, der von dessen Experimenten erfuhr und beschloss, sich an der Paraffineinbettung zu versuchen. His entwässerte Hühnerembryonen in Alkohol, klärte sie mit Lavendelöl und träufelte heißes Paraffin darüber. Dieses Verfahren ermöglichte ihm die Anfertigung guter Schnitte. In der vorliegenden Arbeit haben wir His’ Protokoll der Paraffineinbettung nachvollzogen, um zu ermitteln, ob His tatsächlich die Infiltrationseinbettung mit Paraffin gelungen ist. Wir befolgten das Protokoll aus seiner 1868 erschienenen Monografie. Anhand der Anweisungen von His gelang es uns nicht, Schnitte mit der von ihm illustrierten Qualität herzustellen. In der Regel zerfiel das Gewebe beim Schneiden wegen schlechter Wachsinfiltration. Brauchbare Schnitte wurden nur bei Abwandlung von His’ Protokoll durch das Wiedereinschmelzen der eingebetteten Embryos in frischem Paraffin erreicht. Möglicherweise ist es uns nicht gelungen ist, das Protokoll von His detailgetreu zu replizieren, oder das Protokoll war unvollständig. Nach unserer Einschätzung hat His wahrscheinlich die Infiltrationseinbettung mit Paraffin erfunden und perfektioniert, aber kein vollständiges Protokoll veröffentlicht.

The macro- and microscopic structure of the lungs of mature rabbits – Oryctolagus cuniculus L., 1758 – European rabbit was studied. This work was performed using comprehensive research methods: anatomical, organometric, histological and histometric. The research was carried out in the conditions of the laboratory of pathomorphology in the Department of Anatomy and Histology. The scientific work was performed on the subject of research work of the Department of Anatomy and Histology of Polissya National University on the theme: “Development, morphology and histochemistry of animal organs in normal and pathological”, according to the state registration number – № 0113V000900. The morphological studies followed the basic rules of Good Laboratory Practice GLP (1981), the provisions of the “General Ethical Principles of Animal Experiments”, adopted by the First National Congress of Bioethics (Kyiv, 2001) and the requirements for the “Rules for carrying out work with experimental animals”, approved by the order of the Ministry of Health № 281 of November 1, 2000 “On measures to further improve the organizational forms of work with the use of experimental animals”. The material for histomorphological studies were pieces of lungs selected from clinically healthy adult rabbits (n = 5). Slices were made from paraffin blocks on a sled microtome MS–2. The thickness of histological sections did not exceed 8–12 μm, which were then stained with hematoxylin and eosin and Van Gizon to study the morphofunctional activity of cells and tissues of the respiratory tract. Histometric studies of histological specimens were performed using a microscope “Micros” with a digital camera at a magnification of × 100–1000. Studies have shown that macroscopically the lungs of mature rabbits have a reflection of the shape of the thoracic cavity, and gradually expands ventrally. The right and left lungs are surrounded by pleural right and left sacs. The pleural cavities of both lungs in the test animals are not interconnected. Organometry showed that the absolute lung mass of rabbits is 18.05 ± 1.32 g, relative to 0.624 ± 0.013 %. The histoarchitectonics of the lungs is formed by the lung lobes. The respiratory department of the lungs is represented by a set of acinuses. The latter are represented by respiratory bronchioles, alveolar passages, alveolar sacs and alveoli that form the alveolar tree. The results of morphometric studies showed that the respiratory (respiratory) part of the lungs of rabbits occupies 39.6 ± 0.62 %, and the connective tissue base, respectively, 58.5 ± 1.27 %. The average volume of alveoli, which are divided into small, medium and large, is 42.3 ± 4.35 thousand μm3. Thus, studies of the macro- and microscopic structure of the lungs of experimental animals are of great general biological importance, as they are an important criterion for objective assessment for the differential diagnosis of diseases of various origins.

Objective: The aim of this study was to assess the micro-push-out bond strength of a mineral-based root canal sealer, BioRoot RCS in canals prepared by K3XF rotary systems of two different tapers. Material and Methods: Eighty caries free maxillary central incisors were used in this study. The samples were allocated into 4 groups (n=20) according to the root canal sealer and taper of the rotary instruments. The samples were obturated using single cone obturation technique. From each root 1mm thick slices at coronal, middle and apical thirds were collected using hard tissue microtome under continuous water coolant. Push-out tests were done for these sections using a Universal testing machine (INSTRON 8801) at a crosshead speed of 1mm/min. One-way analysis of variance (ANOVA) was used to compare the bond strengths within groups and Tukey’s multiple post hoc analysis was used for pair-wise comparison of bond strengths. Results: AH Plus exhibited higher micro-push-out bond strength than BioRootRCS though they did not differ significantly (p>0.05). Preparation of root canals with 6% taper rotary instruments showed higher bond strength than 4% though they did not differ significantly (p>0.05). Conclusion: There was no significant difference between micro-push-out bond strength values of BioRoot RCS and AH Plus. The bond strength values were high in 6% taper canals than 4% canals though the difference was not significant statistically.

Understanding the structure and working principle of brain neural networks requires three-dimensional reconstruction of brain tissue samples using array tomography method. In order to improve the reconstruction performance, the sequence of brain sections should be collected with silicon wafers for subsequent electron microscopic imaging. However, the current collection of brain sections based on silicon substrate involve mainly manual collection, which requires the involvement of automation techniques to increase collection efficiency. This paper presents the design of an automatic collection device for brain sections. First, a novel mechanism based on circular silicon substrates is proposed for collection of brain sections; second, an automatic collection system based on microscopic object detection and feedback control strategy is proposed. Experimental results verify the function of the proposed collection device. Three objects (brain section, left baffle, right baffle) can be detected from microscopic images by the proposed detection method. Collection efficiency can be further improved with position feedback of brain sections well. It has been experimentally verified that the proposed device can well fulfill the task of automatic collection of brain sections. With the help of the proposed automatic collection device, human operators can be partially liberated from the tedious manual collection process and collection efficiency can be improved.

Modern studies of detailed processes in plant development would not be possible without using a wide range of fluorescent proteins (R. Day and M. Davidson, 2009; D. Chudakov et al., 2010). However, application of fluorescent proteins is restricted due to problems with their visualization in plant tissues. Plants are difficult objects for microscopic studies: even the most advanced methods have significant limitations regarding the depth of light penetration due to the scattering and absorption of light by cell walls. Therefore, to study the distribution of reporter fluorescent proteins in large organs typical for most plants, it is necessary to fix the plant material and prepare thick histological sections with a vibrating-blade microtome. Chemicals traditionally used for fixing, dehydrating and embedding of plant tissues samples lead to changes in the structure of fluorescent proteins and, as a result, often to the loss of their fluorescence. Therefore, it is important to optimize the protocols for fixing plant tissues, preparing sections and studying the distribution of fluorescent proteins by laser scanning confocal microscopy. In this work, we have proposed for the first time an integrated approach to fixation of tissues of transgenic plants and preparation of sections in the course of the study of patterns of cellular response to auxin and expression of transcription factors using laser scanning confocal microscopy. Our aim was to sum up modern approaches to the application of an effective universal technique for visualization of tissue and cellular patterns of the fluorescent reporter proteins distribution on sections of large non-model plants. The first step for using fluorescent proteins in plants is the generation of genetic constructs that carry the promoter of the gene of interest fused to a reporter gene encoding a fluorescent protein. In this case, a transformation protocol has to be available for the selected plant species. We have described the use of the Gateway® cloning technology for the construction of vectors for plant transformation that meet modern experimental requirements. In order to study auxin localization in vivo we developed a series of vectors with genes encoding various fluorescent proteins (eGFP, tdTomato, mRuby3) under the control of the auxin-sensitive DR5 promoter (E. Ilina et al., 2012). We now demonstrate the advantage of nuclear-targeted fluorescent proteins (mNeonGreen-H2B, tdTomato-H2B, mRuby3-H2B), as well as the possibility of their application for additional visualization of cell nuclei in combination with highly specific cell wall staining using SCRI Renaissance2200. An effective method for constructing vectors to study cell-specific expression patterns of developmental regulators is presented using the transcription factor genes GATA24 (A. Kiryushkin et al., 2019) and LBD16 in some Cucurbitaceae species as examples. We also applied an expression cassette, pAtUBQ10::DsRED1 (E. Limpens et al., 2004), carrying a gene encoding red fluorescent protein under the control of a constitutive promoter, to demonstrate the advantages of the use of fluorescent proteins in screening for transgenic vs. wild type roots. A new method of fixation and clearing of plant tissues containing reporter fluorescent proteins and preparation of sections is presented for the example of transgenic roots of Cucurbitaceae. The advantage of using melted agarose compared to embedding media for orienting plant parts during the preparation of samples was demonstrated. The increase of the photostability of fluorescent proteins in sections due to the use of clearing reagent ClearSee (D. Kurihara et al., 2015) as a mounting medium is demonstrated. Altogether, we present a combination of the advantages that are provided by application of a complex of several modern methodical approaches of sample preparation and laser scanning confocal microscopy that offers significant advantages for studying of the developmental biology of large non-model plants.

Synchrotron radiation-based infrared microspectroscopy (SR-IMS) is a nondestructive bioanalytical technique with a high signal-to-noise ratio and high ultraspatial resolution (3-10 μm). It is capable to explore the microstructures of plant tissues in a chemical sense and provide information on the composition, structure, and distribution of chemical compounds/functional groups. The objective of this study was to illustrate how SR-IMS can be used to image the internal microstructures of chickpea seed tissue within a cellular level. Chickpea seeds (CDC Cory) were collected from the Crop Development Center (University of Saskatchewan, Saskatoon, SK). The seeds were frozen at -20 °C on object disks in a cryostatic microtome and then were cut into thin cross sections (ca. 8 μm thick). The experiment was carried out on the mid-infrared beamline (01B1-1) at the Canadian Light Source (Saskatoon, SK). We obtained the ultraspatial images of the chickpea tissue with pixel-sized increments of imaging steps. The results showed that, with the extremely bright synchrotron light, spectra with high signal-to-noise ratios can be obtained from an area as small as 3.3 μm × 3.3 μm, allowing us to observe the seed tissue within a cellular level. Chemical distribution of chickpea such as lipids, protein, and carbohydrates could be mapped, revealing the chemical information of the chickpea internal microstructure. In conclusion, SR-IMS can rapidly characterize the molecular structure of protein, carbohydrates, and lipids at an ultraspatial resolution.

Evaluation of the biomechanical properties of soft tissues by measuring the stress-strain relationships has been the focus of numerous investigations. The accuracy of stress depends, in part, upon the determination of the cross-sectional area (CSA). However, the complex geometry and pliability of soft tissues, especially ligaments and tendons, make it difficult to obtain accurate CSA, and the development of CSA measurement methods of soft tissues continues. Early attempts to determine the CSA of soft tissues include gravimetric method, geometric approximation technique, area micrometer method, and microtomy technique. Since 1990, a series of new methods have emerged, including medical imaging techniques (e.g. magnetic resonance imaging (MRI), computed tomography (CT), and ultrasound imaging (USI)), laser techniques (e.g. the laser micrometer method, the linear laser scanner (LLS) technique, and the laser reflection system (LRS) method), molding techniques, and three-dimensional (3D) scanning techniques.

The aim of the study is to characterise the cellular structures in tension wood and compare its effect on the selected anatomical parameters in two mangrove wood species viz. Excoecaria agallocha Linn. and Sonneratia caseolaris (L.) Engl. Wood blocks of 1 cm3 were prepared and thin sections of cross sections were cut using microtome and stained with Azure II to differentiate the structure of normal wood from tension wood fibres. The anatomical parameters analysed in this present study were vessel frequency, tangential vessel diameter and fibre wall thickness in tension wood as well as in normal wood. Vessel diameter and vessel frequency were significantly lower in tension wood compared to the normal wood in both the species. The reduction of tangential diameter in tension wood region is a way of protecting these elements from cavitation and providing mechanical support for the plant than liquid transport. A significant variation in fibre wall thickness was observed between tension wood and normal wood in both species.

Objective: The teeth play important roles in food mastication, prehension and defense against predators. Although several studies demonstrated the development of teeth in different mammalian species, no data are, to our knowledge, available in Egyptian buffalos. Therefore, the present investigation was conducted to study the development of incisor teeth in buffaloe. Design: Descriptive study. Animals: seventeen buffalo embryos and fetuses of both sexes were used. Their crown vertebral rump lengths (CVRL) ranged from 2 to 46 cm (equivalent to 38 - 177 days old). Procedure: Embryos and fetuses were fixed in 10% neutral buffered formalin and decalcified by 14% EDTA solution for several weeks. The samples were dehydrated, cleared and embedded in paraffin wax using standard techniques. Sections were cut on Leitz microtome and mounted on uncoated slides. For general histological structure, a selection of slides was routinely stained with haematoxylin–eosin and examined by the light microscope. Result: The present study reported for the first time that the prenatal development of incisors in buffalo passes through three sequential stages: the bud, cap and bell stages. Although each stage was easily differentiated based on its form and its distinct developmental features, the three stages were overlapped with each other’s. Generally, the bud stage was observed in 11 cm CVRL buffalo fetuses, meanwhile the cap and bell stages were detected in 23 cm and 30 cm CVRL fetuses respectively. Conclusion and clinical relevance: These results can be used as inductive index for determination of the ages of buffalo embryos and fetuses.

Objective The aim of this study was to compare the microstructure of blood vessels and nerve fibers between different acupoints in the rat skin from the perspective of histology. Methods The skin tissue at “Qūze” (曲泽 PC3)” “Daling (大陵 PC7)” and “Zhōngchōng (中冲 PC9)” regions was taken from the rat forelimb along the pericardium meridian, and cut sagittally with a freezing microtome. After that, the skin sections were stained by fluorescent histochemistry and immunohistochemistry with phalloidin and calcitonin gene-related peptide (CGRP) to reveal the microstructure of blood vessels and nerve fibers respectively, and then examined under a laser scanning confocal microscope. Results The microstructure of blood vessels and nerve fibers was clearly labeled with phalloidin and CGRP, respectively. The blood vessels and nerve fibers were observed in the dermis of “PC3” “PC7” and “PC9” regions, where the bundle of nerve fibers distribute in parallel to the blood vessels, and sent out thin branches to surround the wall of the blood vessels. In contrast, the density of blood vessels and nerve fibers in “PC3” “PC7” and “PC9” were presented orderly in an increased tendency. Conclusion The results of present study demonstrate the differences of the neurovascular microstructure among “PC3” “PC7” and “PC9” in the rat, providing a histological view to insight into the specificity of different acupoints.

Decalcification of mineralized samples for microscopic analysis involves competing factors including decalcification time, preservation of tissue integrity and cost. We investigated the utility of different decalcification solutions for studying joints in AG/WT, BALB/c, C57, DBA1/J mice and Wistar rats. The hind paws of the rodents were removed and fixed with 10% buffered formalin. Specimens were divided randomly into three groups for demineralization: 10% nitric acid, 12.5% EDTA at room temperature and 12.5% EDTA at 35 °C with shaking. Sections of joints were stained with hematoxylin and eosin (H & E). We evaluated decalcification time and expense, ease of cutting sections, preservation of nuclear basophilia and intranuclear detail, and intensity of eosin staining. The 10% nitric acid solution produced the most rapid decalcification for the mice, but not the rats. The 12.5% EDTA solution at 35 °C with shaking did not decrease decalcification time. Effects on microtomy were variable as were the effects on H & E staining. The EDTA solution provided the best basophilia and intranuclear detail for the mice. For rats, only 12.5% EDTA at 35 °C with shaking produced good preservation. Preservation of nuclear basophilia and intranuclear detail for rats was best with 10% nitric acid and EDTA 35 °C. For mice, 10% nitric acid failed to preserve nuclear basophilia and intranuclear detail. For intensity of eosin staining, EDTA at room temperature and EDTA 35 °C was best for both mice and rats. Sections also exhibited good H & E staining in most samples decalcified with 10% nitric acid. Although we found considerable variation among groups of animals, we found less variation among the different mouse strains than between mice and Wistar rats.

Mast cells, known as pro-inflammatory effector cells, are immunocytes present in the meninges and may be involved in the pathophysiology of migraine. This study aims to evaluate the histomorphometric parameters of mast cells located in the convexity of the human intracranial dura mater. For this, samples of intracranial dura mater from eight human fresh cadavers were collected between 8- and 24-h post-mortem. The whole samples were fixed and, subsequently, two fragments of 1.5 cm² each were cut from four different areas of the dura mater convexity, containing a segment of the middle meningeal artery, totaling 64 fragments. After histological processing, the fragments were submitted to microtomy (5 and 10 µm), stained with toluidine blue (0.1%), or immunohistochemically labeled for tryptase, and analyzed using optical microscopy. The following histomorphometric parameters were evaluated: distance from mast cells to vessels, the density of mast cells, and percentage of mast cells with degranulation. Histomorphometric analyzes showed a higher density of mast cells in the vicinity of blood vessels (arterial and venous), with distances around 0-150 µm. A greater number of mast cells was detected near venous vessels in the periosteal layer (17.0 ± 10.1 cells/mm²) than in the meningeal layer (14.1 ± 7.0 cells/mm²) (p < 0.05). Mast cells from the region close to the superior sagittal sinus were found in greater quantity close to the venous vessels (16.7 ± 10.1 cells/mm²) than to the arterial vessels (11.2 ± 7.5 cells/mm²) (p < 0.05). In short, in the convexity of the human intracranial dura mater, mast cells are located close to blood vessels, with a greater number of cells next to the venous vessels of the periosteal layer and in the proximal region of the superior sagittal sinus.

The objective of this study was to provide optimized processing for examination of rat incisors in nonclinical toxicity studies that enables analysis using immunohistochemistry (IHC). Rat maxillas and mandibles were decalcified in Immunocal, a formic acid decalcifier, and Decal Stat, a hydrochloric acid decalcifier, to evaluate tissue quality when with hematoxylin and eosin (H&E) stain and an IHC. Following necropsy of 10 to 13-week-old male Sprague Dawley rats, tissues were collected, trimmed, fixed in neutral buffered formalin (NBF), and placed into the corresponding decalcifying solution. After a pilot study with multiple timepoints for both decalcifying solutions, times were selected for the definitive study. Incisors in the definitive study were decalcified for 72, 96 or 120 hours in Immunocal and 24 hours in Decal Stat, trimmed, processed, embedded in paraffin, and sectioned. The microtomy process and sections were evaluated by histotechnologists. Sections were stained withH&E or an IHC to detect vimentin. Veterinary pathologists used blinded assessment to evaluate staining and tissue quality. The H&E sections from Immunocal timepoints scored higher based on criteria such as cellular morphology. However, tissue quality decreased at 120 hours with Immunocal but was adequate after 24 hours with Decal Stat. For IHC, moderate to excellent expression of vimentin was observed at timepoints for both decalcifiers. Optimal tissue sectioning and histological quality were achieved on incisor sections decalcified for 96 hours with Immunocal and 24 hours with Decal Stat.

This study aimed to examine the penetration depth into dentinal tubules of some chelating agents. The 17% EDTA and two preparations containing surfactants (Smear Clear, Bioakt Endo) were tested. Surface tension and liquid viscosity were measured using a Dynamic Contact Angle Analyzer and a Haake rotational rheometer. To measure the penetration depth inside dentinal tubules, thirty maxillary central incisors were selected from a pool of extracted human permanent teeth and allocated to three experimental groups (10 samples each), as well as were mechanically shaped and cleansed with 5.25% NaOCl, followed by each of the chelators being labeled with 0.1 wt % Rhodamine B according to final irrigation protocol established. The samples were embedded in an epoxy resin, after which 200 μm thick transverse sections were obtained at 2, 5, and 8 mm from the apex with a saw microtome. The specimens were then observed using a confocal laser microscope (CLSM) and the penetration of the labeled solution was measured in every third of each sample. Statistical analysis was performed using ANOVA or Kruskal–Wallis tests according to the distribution of data, evaluated with the Shapiro–Wilk normality test. Viscosity and surface tension tests have shown that BioAKT Endo has the lowest values compared to EDTA and Smear Clear. The medium penetration depth did not significantly differ among the three irrigants, while it increased considerably from the apical to the coronal level in all groups. Additionally, the maximum penetration depth increased significantly from the apical to coronal level, while among groups, BioAKT Endo showed the highest values at the apical and middle level compared to the other irrigants. No significant differences were observed among the three groups in medium and maximum penetration depths when the entire root was considered. New irrigants containing surfactants show reduced surface tension and, in one case (BioAKT Endo), viscosity. The lowering of the surface tension allows for better penetration of liquids into dentinal tubules than EDTA alone, thus improving the cleaning of the root canal system.

The inferior lobes are prominent bilateral brain areas in the hypothalamus of neopterygians among the ray-finned fishes. They are known as multisensory integration centers. As such, they should play a major role in fish evolution. In this study, a comparative morphometric analysis was performed. The morphology of the hypothalamus, where the inferior lobe is considered as fully developed first in Lepisosteus, was then re-examined. One hundred brains from different species of 60 families of ray-finned fishes were stained with cresyl violet and embedded in methacrylate. They were then cut on a microtome while conducting block-face imaging. The volumes were determined for the whole brain, brain areas, and nuclei. Since visual input represents a major sensory input for the inferior lobe, the nucleus glomerulosus, a visual-related nucleus in paracanthopterygian and acanthopterygian teleosts, and the tectum opticum were included in the investigations. The morphometric analysis revealed that the relative volume of the inferior lobes increases significantly from species of the Lepisosteiformes to the Tetraodontiformes. In addition, a positive correlation was detected between the relative volume of the inferior lobes and either the relative volume of the nucleus glomerulosus or the relative volume of the tectum opticum. These correlations, in combination with findings from previous hodological and behavioral studies, give rise to the speculation that the inferior lobes may be involved in higher cognitive processes and complex social interactions.

Currently two techniques exist for 3D reconstruction of biological samples by time-of-flight secondary ion mass spectrometry (ToF-SIMS). The first, based on microtomy and combining of successive section images, is successfully applied for tissues, while the second, based on sputter depth profiling, is widely used for cells. In the present work, we report the first successful adaptation of sectioning technique for ToF-SIMS 3D imaging of a single cell-fully grown mouse germinal vesicle (GV) oocyte. In addition, microtomy was combined with sputter depth profiling of individual flat sections for three-dimensional reconstruction of intracellular organelles. GV oocyte sectioning allowed us to obtain molecule-specific 3D maps free from artifacts associated with surface topography and uneven etching depth. Sputter depth profiling of individual flat slices revealed fine structure of specific organelles inside the oocyte. Different oocyte organelles (cytoplasm, germinal vesicle, membranes, cumulus cells) were presented on the ion images. Atypical nucleoli referred to as "nucleolus-like body" (NLB) was detected inside the germinal vesicle in PO3- and CN- ions generated by nucleic acids and proteins respectively. Significant difference in PO3- intensity in the NLB central area and NLB border was found. This difference appears as a bright halo around the center area. The NLB size calculated for PO3- and CN- ion images is 12.9 ± 0.2 μm and 11.9 ± 0.2 μm respectively, which suggests that bright halo of PO3- ions is a chromatin compaction on the NLB surface. Areas of approximately 1.0-2.5 μm size inside nucleoplasm with increased PO3- and CN- signal were registered in germinal vesicle. Observed compartments have different sizes and shapes, and they are likely attributed to chromocenters or chromosomes.