ALBERT

Description: ABSTRACT The objective of this study was to analyze the dentin-resin cements interfacial ultramorphologies using two different methods: scanning (SEM) and transmission electron microscopy (TEM). Four commercial products were evaluated: two conventional cementing system (RelyX ARC/Adper™ Scotchbond™ Multi-Purpose Plus, 3M ESPE and Clearfil Esthetic Cement/DC Bond, Kuraray) and two self-adhesive resin cements (RelyX Unicem, 3M ESPE and Clearfil SA Cement, Kuraray). Prepolymerized resin disks (Sinfony, 3M ESPE) were cemented on oclusal dentin surfaces of 24 third human molars, simulating the indirect restorations. After 24 h, teeth were sectioned into 0.9-mm thick slabs and processed for microscopy analyses (SEM or TEM/ n = 3). Qualitative characterization of dentin-resin cement interface was performed. Hybrid layer formation with long and dense resin tags was observed only for RelyX ARC cementing system. Clearfil Esthetic Cement/DC Bond system revealed few and short resin tags formation, whereas no hybridization and resin tags were detected for self-adhesive resin cements. Some interfacial regions exhibited that the self-adhesive resin cements were not bonded to dentin, presenting bubbles or voids at the interfaces. In conclusion, TEM and SEM bonding interface analyses showed ultramorphological variations among resin cements, which are directly related to dental bonding strategies used for each resin cement tested. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT Mimosa pudica has three distinct specialized organs, namely, pulvinus, secondary pulvinus, and pulvinule, which are respectively controlling the movements of petioles, leaflets, and pinna in response to external stimuli. Water flow is a key factor for such movements, but detailed studies on the organization of the vascular system for water transport in these organs have not been published yet. In this study, organizations of the xylem vessels and morphological features of the pulvinus, the secondary pulvinus, and the pulvinule were experimentally investigated by X-ray computed tomography and histological technique. Results showed that the xylem vessels were circularly distributed in the specialized motile organs and reorganized into distinct vascular bundles at the extremities. The number and the total cross-sectional area of the xylem vessels were increased inside the specialized motile organs. Morphological characteristics obtained in this study provided new insight to understand the functions of the vascular networks in the dynamic movements of M. pudica . Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT Estimates of the radii and solute concentrations of simulated microstructures containing ultrafine spherical precipitates were determined from isoconcentration surfaces and proximity histograms. The accuracy of the estimates of the solute concentrations and the radii of precipitates was found to depend on the size of precipitates. Optimized parameters for analyzing 0.5- to 2-nm-radius precipitates are proposed. The solute content of 0.5-nm-radius precipitates was not estimated correctly by this method. The accuracy of the estimates of the solute concentration and the radius of precipitates were primarily influenced by the solute concentrations of the precipitates. The ranges of error of the solute concentration in the precipitates, which are associated with the analytical limitations of the ultrafine precipitates, were determined, and the results indicated a limitation of the estimates. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT Ovate mitochondria in cultured human fibroblasts divide by pinching. In the process, as observed by transmission electron microscopy, a deep incisure of the surface membranes separates the organelle into two lobes connected by a slender isthmus. A single element of smooth endoplasmic reticulum (SER) invariably accompanies each incisure, extending deep into the cleft. When the ingrowing membranes meet and fuse with the antipodal membranes, fission occurs. Elongated mitochondria that give no indication of division often are cloaked by a single, continuous cistern of SER. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT The regeneration of axons after a spinal cord injury or disease is attracting a significant amount of interest among researchers. Being able to assess these axons in terms of morphology, length and origin is essential to our understanding of the regeneration process. Recently, two specific axon tracers have gained much recognition; biotinylated dextran amine (BDA) 10 kDa as an anterograde tracer and cholera toxin-B as a retrograde tracer. However, there are still several complexities when using these tracers, including the volume that should be administered and the best administration site so that a significant amount of axons are labeled in the area of interest. In this article, we describe some simple procedures for injecting the tracers and detecting them. We also quantified the number of axons at different locations of the spinal cord. Our results show axons labeled from motor cortex injections traveled down to the lumbosacral spinal cord in 2 weeks, while BDA injections into the lateral vestibular nucleus and reticular formation took 3 weeks to label axons in the lumbosacral spinal cord. Moreover, this protocol outlines some basic procedures that could be used in any laboratory and gives insight into the number of axons labeled and how procedures could be tailored to meet specific researcher's needs. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT The usefulness of embedment-free section transmission electron microscopy (TEM) is stressed for present and future morphological analyses, and several examples are demonstrated which are revealed in sections for the first time by this method: en-face views of slit diaphragm of renal glomerulus and fenestrated diaphragm of capillary endothelium, transparency of neural myelin, attenuated endothelium and some basement laminae, labyrinth architecture of vacuoles within lipid droplets, and enhanced 3D effect of ultrastructures, the latter of which is the case in electron tomography. In addition, the biological significance of structured appearance (microtrabecular lattices) of the cytoplasmic matrix, which is disclosed by this method, are briefly reviewed in relation to the sol–gel transition of cytoplasmic heterogenous proteins. Since the ultrastructures of various cells and tissues in this method are confirmed to be well correspondent to those in conventional epoxy section TEM except for isotropic dimensional changes, and because there is no necessity for any special expensive equipments other than those for the conventional TEM, the embedment-free section TEM method with these advantages, deserves much more wide application to the morphological research including electron tomography. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT Combined microscopy techniques are used to establish the usability of phosphonic acid layers as promoters of hydroxyapatite (HAp) growth. Using spread coating, octadecylphosphonic acid (OPA) self-assembled bilayers are delivered to the thin natural oxide layer of a titanium film surface with no prior treatment. These bilayers aggregate two major advantages of phosphonic moieties to titanium surfaces: nucleation of hydroxyapatite crystals from ionic solution and affinity for both titanium oxide surface and HAp crystals. The functionalized substrates and bare titanium (control) samples are immersed in an aqueous solution containing calcium and phosphorus ions. Over a 4-week immersion time, OPA-functionalized substrates present numerous large agglomerates of inorganic crystals, in contrast to control samples, with no significant amount of deposits. Initial sample characterization was performed with atomic force microscopy (AFM). Compositional and structural characterization of these agglomerates (using TEM, EDS, and electron diffraction), revealed that they are indeed HAp, the main component of the inorganic bone matrix. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT This study focused on adhesive interface morphologic characterization and nanoleakage expression of resin cements bonded to human dentin pretreated with 1% chlorhexidine (CHX). Thirty-two non-carious human third molars were ground flat to expose superficial dentin. Resin composite blocks were luted to the exposed dentin using one conventional (RelyX ARC) and one self-adhesive resin cement (RelyX U100), with/without CHX pretreatment. Four groups ( n  = 8) were obtained: control groups (ARC and U100); experimental groups (ARC/CHX and U100/CHX) were pretreated with 1% CHX prior to the luting process. After storage in water for 24 h, the bonded teeth were sectioned into 0.9 × 0.9 mm 2 sticks producing a minimum of 12 sticks per tooth. Four sticks from each tooth were prepared for hybrid layer evaluation by scanning electron microscope analysis. The remaining sticks were immersed in silver nitrate for 24 h for either nanoleakage evaluation along the bonded interfaces or after rupture. Nanoleakage samples were carbon coated and examined using backscattered electron mode. Well-established hybrid layers were observed in the groups luted with RelyX ARC. Nanoleakage evaluation revealed increase nanoleakage in groups treated with CHX for both resin cements. Group U100/CHX exhibited the most pronouncing nanoleakage expression along with porous zones adjacent to the CHX pretreated dentin. The results suggest a possible incompatibility between CHX and RelyX U100 that raises the concern that the use of CHX with self-adhesive cements may adversely affect resin-dentin bond. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT Owing to the highly efficient two-photon fluorescence of gold nanorods and very short fluorescence lifetime compared with the rotational correlation time, the rotation and diffusion of a single gold nanorod can be easily observed by two-photon fluorescence correlation spectroscopy (TP-FCS). This property, along with the previous successful use as a contrast agent in two-photon fluorescence imaging, suggests a potential application in TP-FCS as well. Although the FCS measurement becomes highly efficient with gold nanorods as probes, the amplitude and temporal decay of the measured correlation functions depend critically on excitation power. Here, we investigate various photophysical processes of gold nanorods to determine the cause of such a sensitive power dependency. This understanding provides a basis for choosing appropriate FCS models to recover reasonable physical parameters. Although the correlation function amplitude G (0) is 32 times lower when the excitation power increases from 20 µW to 1.12 mW, the application of a saturation-modified FCS model yields very good fit to each data set and the fitted concentration of 0.64 nM is comparable to the 0.7 nM given by the inductively coupled plasma mass spectrometry measurement. The FCS assay appears to be an efficient method for the quantification of gold nanorods when correctly interpreted. However, even with the saturation considered in the fitting model, the fitted rotational and translational diffusion rates are getting faster as the power increases. This indicates that other effects such as photothermal effects may raise the local temperature, and thus increasing the rotational and translational diffusion rate. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT Wistar rats (male) were daily administered chlorpyrifos at a dose of 5 mg/kg b wt. and 10 mg/kg b wt. and sacrificed on 1st, 2nd, 4th, 6th, and 8th week. In chlorpyrifos exposed rats hypocalcemia, hypophosphatemia and hypomagnesemia were recorded. At later intervals an increased levels of serum calcium and phosphate were observed. The parathyroid glands and calcitonin cells exhibited increased activity which is evident by increased nuclear volume of these cells. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT In the dry northern temperate regions of the northern hemisphere, the genus Ephedra comprises a series of native shrub species with a cumulative application history reaching back well over 2,000 years for the treatment of asthma, cold, fever, as well as many respiratory system diseases, especially in China. There are ethnological and philological evidences of Ephedra worship and utilization in many Eurasia Steppe cultures. However, no scientifically verifiable, ancient physical proof has yet been provided for any species in this genus. This study reports the palaeobotanical finding of Ephedra twigs discovered from burials of the Gumugou archaeological site, and ancient community graveyard, dated around 3800 BP, in Lop Nor region of northwestern China. The macro-remains were first examined by scanning electron microscope (SEM) and then by gas chromatography-mass spectrometry (GC-MS) for traits of residual biomarkers under the reference of modern Ephedra samples. The GC-MS result of chemical analysis presents the existence of Ephedra -featured compounds, several of which, including benzaldehyde, tetramethyl-pyrazine, and phenmetrazine, are found in the chromatograph of both the ancient and modern sample. These results confirm that the discovered plant remains are Ephedra twigs. Although there is no direct archaeological evidence for the indication of medicinal use of this Ephedra , the unified burial deposit in which the Ephedra was discovered is a strong indication of the religious and medicinal awareness of the human inhabitants of Gumugou towards this plant. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT Objective : This in vitro study evaluated the effect of different endodontic auxiliary chemical substances over Enterococcus faecalis (Ef) biofilm through confocal laser scanning microscopy (CLSM). Methods : Forty-five bovine incisors were infected with Ef for 21 days. Teeth were divided into five groups: group 1: 2.5% NaOCl + EDTA, group 2: 2% CHX gel + EDTA, group 3: 2% CHX liquid + EDTA, group 4: 2.5% NaOCl + 2% CHX gel + EDTA, group 5: 2.5% NaOCl + 2% CHX liquid + EDTA and a negative and a positive control group (NCG; PCG). The samples were stained with SYTO9 and propidium iodide and analyzed by CLSM. Bacterial viability was quantitatively analyzed by the proportions of dead and live bacteria in the biofilm remnants. Scores were standardized according to the total bacterial load (TBL)—1: ≤25%, 2: 〉25 ≤50%, 3: 〉50 ≤75%, 4: 〉75% and debris—1: absence of debris; 2: presence of debris. Statistical analysis was carried out through the Kruskal–Wallis and the Fischer exact tests ( P  = 0.05). Results : No statistical differences were observed to CFU, debris and bacterial viability. Conclusion : None of the tested substances could completely eliminate Ef from the root canal space. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT The aim of the present research was to analyze ultrastructural and immunohistochemical aspects of the alveolar repair after the extraction of molars of alendronate (ALN)-treated rats. Wistar rats received 2.5mg/kg body wt/day of ALN during 14 days previously and 7, 14 and 21 days after the extraction of the second mandibular molar. Specimens were fixed in 2% glutaraldehyde + 2.5% formaldehyde under microwave irradiation, decalcified in 4.13% EDTA and paraffin embedded for TRAP histochemistry and immunohistochemistry for OPN, BSP and endoglin, or embedded in Spurr epoxy resin for TEM analysis. Additional specimens had their soft tissues removed and were processed for scanning electron microscopy. The ALN group presented latent TRAP-positive osteoclasts and nonresorbed alveolar crests with bacterial infection. Mild bone necrosis signs were observed at all time points studied. Ultrastructurally, empty osteocyte lacunae were observed and bone trabeculae surface presented hyalinized aspect. A significant delay in alveolar repair occurred, as well as decreased angiogenesis. ALN treatment provoked mild signs of bone necrosis, despite the high dose employed. The present findings add new information about the ultrastructural aspect of the early repair of rats under ALN treatment and highlight for giving attention when oral surgeries are performed in patients using this drug. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT In the long run, the widespread use of slide scanners by pathologists requires an adaptation of teaching methods in histology and cytology in order to target these new possibilities of image processing and presentation via the internet. Accordingly, we were looking for a tool with the possibility to teach microscopic anatomy, histology, and cytology of tissue samples which would be able to combine image data from light and electron microscopes independently of microscope suppliers. With the example of a section through the villus of jejunum, we describe here how to process image data from light and electron microscopes in order to get one image-stack which allows a correlation of structures from the microscopic anatomic to the cytological level. With commercially available image-presentation software that we adapted to our needs, we present here a platform which allows for the presentation of this new but also of older material independently of microscope suppliers. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT This article presents a pixellated solid-state photon detector designed specifically to improve certain aspects of the existing Everhart–Thornley detector. The photon detector was constructed and fabricated in an Austriamicrosystems 0.35 µm complementary metal-oxide-semiconductor process technology. This integrated circuit consists of an array of high-responsivity photodiodes coupled to corresponding low-noise transimpedance amplifiers, a selector-combiner circuit and a variable-gain postamplifier. Simulated and experimental results show that the photon detector can achieve a maximum transimpedance gain of 170 dBΩ and minimum bandwidth of 3.6 MHz. It is able to detect signals with optical power as low as 10 nW and produces a minimum signal-to-noise ratio (SNR) of 24 dB regardless of gain configuration. The detector has been proven to be able to effectively select and combine signals from different pixels. The key advantages of this detector are smaller dimensions, higher cost effectiveness, lower voltage and power requirements and better integration. The photon detector supports pixel-selection configurability which may improve overall SNR and also potentially generate images for different analyses. This work has contributed to the future research of system-level integration of a pixellated solid-state detector for secondary electron detection in the scanning electron microscope. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT Birds occupy a prominent place in the Brazilian economy not only in the poultry industry but also as an animal model in many areas of scientific research. Thus the aim of this study was to provide a description of macro and microscopic aspects of the ectoderm-derived structures in chicken embryos / fetuses poultry ( Gallus gallus domesticus ) from 1st to 19th day of incubation. 40 fertilized eggs, from a strain of domestic chickens, with an incubation period of 2–19 days were subjected to macroscopic description, biometrics, light, and scanning microscopy. All changes observed during the development were described. The nervous system, skin and appendages and organs related to vision and hearing began to be identified, both macro and microscopically, from the second day of incubation. The vesicles from the primitive central nervous system—forebrain, midbrain, and hindbrain—were identified on the third day of incubation. On the sixth day of incubation, there was a clear vascularization of the skin. The optic vesicle was first observed fourth day of development and on the fifth day there was the beginning of the lens formation. Although embryonic development is influenced by animal line as well as external factors such as incubation temperature, this paper provides a chronological description for chicken ( Gallus gallus domesticus ) during its embryonic development. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT The dog ( canis lupus familiaris ) is the only other species besides humans that develop spontaneous prostatic carcinomas (PCa) at a high frequency. The canine model is primarily utilized for the study of the PCa molecular mechanisms and provides a natural animal model for the study of potential therapies. In humans, the PCa frequently exhibits mutations in the C-MYC and a reduced expression of the E-cadherin and NKX3.1 proteins. This study's objective was to evaluate the NKX3.1, C-MYC, and E-cadherin expression in the canine normal prostate, benign prostatic hyperplasia (BPH), proliferative inflammatory atrophy (PIA) and PCa and to verify differences in expression and subcellular localization of these proteins in the prostatic carcinogenesis. A tissue microarray (TMA) slide was constructed, and immunohistochemistry with antibodies raised against C-MYC, NKX3.1, E-cadherin and p63 was performed using the peroxidase and DAB methods. The C-MYC protein expression was elevated in the cytoplasm and nuclei of the canine PCa and PIA compared with the normal prostate ( P  = 0.004. The NKX3.1 protein expression was reduced in 94.75% of the PCa and 100% of the PIA compared with the normal prostate ( P  = 0.0022). In fact, the expression of E-cadherin trended towards a decrease in carcinomas when compared to normal prostate and PIA. By immunohistochemistry, more p63-positive basal cells were observed in the PCa and PIA when compared with the normal prostate ( P  = 0.0002). This study has demonstrated that the carcinogenesis of canine prostatic tissue may be related to basal cell proliferation, the gain of C-MYC function and the loss of NKX3.1 protein expression. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT Early osteoporosis diagnosis is of important significance for reducing fracture risk. Image analysis provides a new perspective for noninvasive diagnosis in recent years. In this article, we propose a novel method based on machine-learning method performed on micro-CT images todiagnose osteoporosis. The aim of this work is to find a way to more effectively and accurately diagnose osteoporosis on which many methods have been proposed and practiced. In this method, in contrast to the previously proposed methods in which features are analyzed individually, several features are combined to build a classifier for distinguishing osteoporosis group and normal group. Twelve features consisting of two groups are involved in our research, including bone volume/total volume (BV/TV), bone surface/bone volume (BS/BV), trabecular number (Tb.N), obtained from the software of micro-CT, and other four features from volumetric topological analysis (VTA). Support vector machine (SVM) method and k -nearest neighbor ( k NN) method are introduced to create classifiers with these features due to their excellent performances on classification. In the experiment, 200 micro-CT images are used in which half are from osteoporosis patients and the rest are from normal people. The performance of the obtained classifiers is evaluated by precision, recall, and F-measure. The best performance with precision of 100%, recall of 100%, and F-measure of 100% is acquired when all the features are included. The satisfying result demonstrates that SVM and k NN are effective for diagnosing osteoporosis with micro-CT images. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT This study sought to evaluate the efficacy of passive ultrasonic irrigation (PUI) on removing the smear layer and debris from root dentin using scanning electron microscopy (SEM). Twenty-five bovine incisors were manually prepared and divided into three groups according to the final irrigation protocol: EDTA, final irrigation with 12 mL of 17% EDTA for 3 minutes followed by 5 mL of 2.5% NaOCl; EDTA/PUI, final flush with 4 mL of 17% EDTA and PUI for 30 seconds. These procedures were repeated three times to standardize the volume of the irrigant. Control group, after preparation, the specimens were irrigated only with 17 mL of 2.5% NaOCl. The roots were fractured and analyzed using SEM. The intragroup analysis revealed that the EDTA/PUI protocol removed a higher amount of debris at the cervical third ( P  = 0.03). The intergroup analysis revealed that EDTA/PUI presented the lowest amount of debris at the cervical third ( P  = 0.007). Smear layer scores were higher in the control group compared with the EDTA and EDTA/PUI groups, but only at the cervical third ( P  = 0.02). None of the final irrigant protocols completely removed the smear layer and debris. EDTA/PUI only improved the removal of debris at the cervical third. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT The bullfrog ( Lithobates catesbeianus ) is an exotic animal that adapted perfectly to the climatic conditions of Brazil after introduction in the country. These climatic conditions favor the reproduction and fattening of bullfrogs. However, the development of the bullfrog is significantly reduced in colder regions because low temperatures affect the secretion of gonadotropins and change the sensitivity of the germinal epithelium to gonadotropic hormones. Thus, the temperature and photoperiod are abiotic factors that influence the life cycle of this species, especially with regard to reproduction. Samples from different regions of the oviducts (the infundibulum, magnum, and isthmus) of 9 bullfrogs were collected in the different seasons of the year. The samples were fixed and processed for analysis with scanning and transmission electron microscopy. A comparison of the morphology of the oviducts collected during different seasonal periods suggests that the frogs have greater reproductive potential in the spring due to the development of the structures related to the oviduct, presence of developed cilia, and electron-dense granules. However, seasonal changes were noted in the animal throughout the year in preparation for reproduction during the spring and summer. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT Background : Cancellous bone defects surrounded by still intact bone structures never heal. Ceramics offer a solution providing osteoconductive scaffolds. Purpose : The purpose of the study is to evaluate whether structured β-TCP and HA implants can reconstruct cancellous bone defects, which role micro- and macro-porosity, stiffness and surface area play; finally the indication for both materials based on its resorbability. Material & Methods : 10 German Shepard dogs were operated on both tibial heads implanting shell-like fully interconnected ceramic cylinders, using a wet grinding hollow drill coated with diamonds. β-TCP was compared with HA. A polychromatic sequential labelling with 4 different fluorochromes controlled bone formation dynamics. Non-decalcifying histology after perfusion fixation and vessel casting was performed. μ-CT was combined with high resolution microradiography and histology on thin ground crossections. The stages after 6 weeks, 2, 3, 4 months and 15 months were evaluated. Results : In spite of osseointegration of HA and β-TCP, the osseointegration of both materials was completely different. Both shell-like bone void fillers were osseointegrated in a sandwich-like manner. HA yielded primarily a reinforcement of the recipient's cancellous-bone bed and full osseointegration after 4 months, whereas β-TCP-implants were fully osseointegrated after 6 weeks. HA did not show signs of resorption. The resorption of the β-TCP resulted during remodelling. The final stage showed restitution “ad integrum” of the β-TCP defects with a physiological architecture, whereas HA was integrated in the cancellous bone construction providing 600 μm measuring macropores showing osteoinductive properties. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT The structural and morphological properties of the teratoma membrane were investigated to better understand the pathogenesis of ovarian teratomas. A mature cystic teratoma and amnion were obtained from patients who underwent laparoscopic cystectomy and uncomplicated delivery, respectively. The teratoma membrane was divided into three layers according to the results of the histological analysis. Each layer showed distinct morphological properties, including an outer layer that was uniformly arranged, a middle layer with an irregular pattern of fibers, and an inner layer that was structurally dense with a wavy pattern of fibers. The morphology of the layers of the amniotic membrane was the reverse that of the teratoma membrane. In the teratoma membrane, the outer layer was primarily composed of type III collagen and the inner layer had a large amount of type III and IV collagen. The amniotic membrane showed a small amount of type III collagen in the outer layer, whereas the inner layer had large amounts of type I, III, and IV collagen. In the teratoma membrane, the collagen fibrils were arranged regularly in the outer layer, but irregularly in the inner layer. In the amniotic membrane, the arrangement of collagen fibrils was the reverse that of the teratoma membrane. Additionally, the collagen fibrils in the teratoma membrane were thinner than those of the amniotic membrane and had slightly shorter d -spacing. Two membranes showed the differences in collagen fibril arrangement, which may caused by the different functional roles. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT The rock cavies ( Kerodon rupestris ) are rodents belonging to family Caviidae, with habitat restricted to northeastern Brazil. Current studies have shown that blood cord has stem and hematopoietic cells both with a high regenerative potential, microscopic studies about the description in rodents and other mammals are scarce. The aim of this study was to describe the structural components of the umbilical cord of rock cavies. Rock cavies embryos at different stages of pregnancy were used, three in the first third, five in middle third, and six in the final third of gestation. They were obtained at the Centre for Wild Animals Multiplication of Federal Rural University of Semi-Arid. In rock cavies was observed that the number of vessels involved in the transport of maternal substances was represented by two major arteries, veins and an allantois duct. The umbilical cord was delimited by amniotic epithelium, formed by a layer of squamous cells, connective tissue with characteristic of the mucosal tissue, corresponding to the Wharton jelly (fetal mesenchyme), where elastic fibers were observed and intercellular spaces were arranged in concentric blades. The morphology observed in rock cavies is similar to that described in the literature for other mammals, including man. The rock cavies are well adapted to captivity and are docile, which makes this animal an important model for study within the field of regenerative medicine. The knowledge of the umbilical cord morphology represents a base point for using this animal as a model for culture and cell therapy. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT Semi-oriented microstructures represent an intermediate state between isotropic and complex oriented microstructures of serpulids. Semi-oriented microstructures are apparently unique to serpulids, indicating that there may have been unique aspects in the evolution of their biomineralization system. The occurrence of semi-oriented microstructures in serpulids supports the hypothesis that complex oriented microstructures are derived from isotropic microstructures via evolution of a biochemical crystal orientation control mechanism. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT The aim of this study was to evaluate, using atomic force microscopy, the effect of two different bleaching agents on the modification of dental composites materials. This modification will be judged by analyzing the variation of surface roughness and surface morphology of two different composites: one containing nanoparticles and other consisting of microhybrid resin. The bleaching was performed by using two different concentrations of hydrogen peroxide: HP Blue (20% hydrogen peroxide) and Whiteness HP Maxx (35% hydrogen peroxide). Disks of Esthet X and Filtek™ Z350 composites were used. Atomic force microscopy was used for analyses of the same place of the sample before and after treatment. A total of 12 analyses were performed per group ( n  = 12). The samples were analyzed qualitatively by evaluating morphological changes in the images and quantitatively by using roughness parameters (Ra). Data were analyzed statistically using Kruskal–Wallis, Mann–Whitney, and Friedman tests ( P  〈 0.05). Changes were observed both qualitatively and quantitatively only in the groups where Esthet X resin was used. The use of hydrogen peroxide bleaching agents caused changes only in the surface of microhybrid composites, with no changes being observed in the composite containing nanoparticles. Despite being even significant, these alterations are clinically slight and can be eliminated by polishing them. Microsc. Res. Tech., 2013. © 2013 Wiley Periodicals, Inc.

Description: Peanut agglutinin (PNA) and Ulex europaeus agglutinin-1 (UEA-1) were used as probes to study the distribution of β-gal (1→3) ga1Nac and α- l -Fucose in rabbit uterus during early pregnancy. PNA binding was mainly localized on the surface of uterine glandular and luminal epithelium. There were no positive signals on day 1 of pregnancy. PNA binding gradually increased from day 2 and reached its highest level on days 3 and 4. The distribution of PNA binding gradually declined from day 5 and reached a low level on day 7. However, UEA-1 binding was only localized on the luminal epithelial during early pregnancy. A high level of UEA-1 binding had been found on the luminal epithelium on day 1 of pregnancy and low level of positive signals had been found in the uterus on days 2 and 3. UEA-1 binding increased gradually and reached its highest level on day 4. Then the distribution of UEA-1 binding sharply declined and no positive signals were found on days 5–7. The distribution of PNA and UEA-1 bindings in pseudopregnant uterus was similar to that in normal pregnant uterus. During estrus cycle, there was no detectable PNA binding signal in uterus. But, a high level of UEA-1 binding was found in the luminal epithelium of estrus uterus. In ovariectomized rabbit uterus, progesterone significantly induced the expression of PNA binding, while estrogen stimulated UEA-1 binding expression. These results suggested that the distribution of PNA and UEA-1 bindings in rabbit uterus may be related to rabbit implantation. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT The embryonic origin of the alimentary canal, especially the midgut, is a controversial problem in insects, and it has not been satisfactorily resolved to date. The organogenesis of the digestive system in the embryonic development was observed in the scorpionfly Panorpa obtusa Cheng using light, transmission, and scanning electron microscopy. The embryonic development lasts about 150–160 h at 24°C. The stomodaeum is formed from an invagination in the medioposterior portion of the protocephalon mid-ventrally posterior to the labral segment at 76 h after oviposition. The proctodaeum arises as an invagination from the caudal end of the abdomen at 78 h. Four anal forks are formed from within the opening of proctodaeum. Three pairs of proctodaeal evaginations are formed from the anterior part of the proctodaeum, and eventually developing into Malpighian tubules, thus are of ectodermal origin. The cardiac and pyloric valves develop from stomodaeum and proctodaeum, respectively, and also of ectodermal origin. The midgut epithelium originates from anterior and posterior midgut rudiments in blind ends of the stomodaeum and proctodaeum, and it is of endodermal origin. The two cell-bands (rudiments) cover the yolk ventrally and then dorsally, elongate to each other, and eventually fuse to form the midgut. The midgut formation pattern is briefly discussed in different insects. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT Recently, atomic force microscope (AFM) manufacturers have begun producing instruments specifically designed to image biological specimens. In most instances, they are integrated with an inverted optical microscope, which permits concurrent optical and AFM imaging. An important component of the set-up is the imaging chamber, whose design determines the nature of the experiments that can be conducted. Many different imaging chamber designs are available, usually designed to optimize a single parameter, such as the dimensions of the substrate or the volume of fluid that can be used throughout the experiment. In this report, we present a universal fluid cell, which simultaneously optimizes all of the parameters that are important for the imaging of biological specimens in the AFM. This novel imaging chamber has been successfully tested using mammalian, plant, and microbial cells. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: The development of a novel three-dimensional image analysis technique of stacks generated by confocal laser scanning microscopy is described allowing visualization of mitochondria-rich cells (MRCs) in the seawater-adapted Nile tilapia in relation to their spatial location. This method permits the assessment and classification of both active and nonactive MRCs based on the distance of the top of the immunopositive cell from the epithelial surface. In addition, this technique offers the potential for informative and quantitative studies, for example, densitometric and morphometric measurements based on MRC functional state. Confocal scanning laser microscopy used with triple staining whole-mount immunohistochemistry was used to detect integumental MRCs in the yolk-sac larvae tail of the Nile tilapia following transfer from freshwater to elevated salinities, that is, 12.5 and 20 ppt. Mean active MRC volume was always significantly larger and displayed a greater staining intensity (GLM; P 〈0.05) than nonactive MRCs. Following transfer, the percentage of active MRCs was seen to increase as did MRC volume (GLM; P 〈0.05). Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT Objective : The objective of this study was to describe the microstructure of different types of gallbladder stones to better understand the basis of gallbladder stone formation. Methods : Gallbladder stones from 387 patients with cholecystolithiasis were first analyzed by Fourier transform Infrared spectroscopy to identify the type of the gallbladder stone, and they were then examined using scanning electron microscopy to define their microstructure. Results : Cholesterol stones were mainly composed of plate-like or lamellar cholesterol crystals stacked tightly in a radial, cord-like, or irregular staggered arrangement. A small number of bilirubinate particles were seen occasionally. Pigment stones were mainly composed of loosely arranged bilirubinate particles with different shapes (sphere-like, clumping-like, or amorphous). Calcium carbonate stones were composed of calcium carbonate crystals having many shapes (bulbiform, ellipsoid, fagot-shaped, fusiform, hawthorn-shaped, cuboid, button-shaped, lamellar, broken firewood-shaped, rod-shaped, acicular, or crushed sugar cane-shaped). Bulbiform shaped crystals were the most common. Phosphate stones were mainly composed of different sized echin-sphere-like or rough bulbiform crystals. Bilirubinate particles were always adherent. Calcium stearate stones usually had a network structure with adherent bilirubinate particles. Protein stones usually had a honeycomb or chrysanthemum petal-like structure. Cystine stones were composed of hexagonal cystine crystals, some of which had prominent edges. Mixed stones presented different shapes according to their mixed components. Conclusion : Different types of gallbladder stones had characteristic shapes and elements. This study provides an objective basis for further research regarding gallbladder stone formation. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT Chrysolina aeruginosa Fald. (Coleoptera, Chrysomelidae) is an important pest of Artemisia ordosica Krasch. In recent years, this phytophagous beetle has spread rapidly throughout northwest China, which has led to mass mortalities of the A. ordosica . This pest has caused great damage to the local ecology. To address this problem, this study compared the sensilla types and their densities on the antennae, maxillary palps, and labial palps of C. aeruginosa . Six different types of sensilla were observed on the antennae using scanning electron microscopy, i.e., sensilla chaetica, sensilla trichodea, sensilla basiconica, sensilla coeloconica, sensilla styloconica, and Böhm bristles. Sensilla chaetica were the main sensilla on the antennae. There was a significant difference in the distribution and density of sensilla trichodea between males and females. Sensilla basiconica were distributed from the fifth to the ninth flagellomeres. Sensilla coeloconica and sensilla styloconica were usually found from the fifth to the eighth flagellomeres. Böhm bristles were found only on the terminal region of the scape and pedicel. Sensilla chaetica and sensilla twig basiconica were observed on the maxillary and labial palps. Sensilla chaetica were distributed all over the maxillary and labial palps. Sensilla twig basiconica were located in the distal areas of these two types of palps. In this study, we also discussed the responses of C. aeruginosa to the volatile semiochemicals produced by their host plants and the behaviours exhibited during host and habitat selection. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT Objective To check if ovariectomy produces alterations in the articular cartilage structure of the tibial proximal epiphysis and if aerobic physical exercise has any influence on these alterations. Methods Six-month-old female Wistar rats were distributed in three groups: Control group (CG), without ovariectomy and physical activity; Oophorectomy and sedentary group (OSG), with bilateral ovariectomy and without physical activity, and Oophorectomy and trained group (OTG), with bilateral ovariectomy and physical activity during 3 months. All rats were euthanised at 9 months of age. The results were obtained by using histological sections stained with Hematoxylin and Eosin, and Picrosirius stain. Results In relation to the thickness of the articular cartilage an increase was observed in the superficial layer in OSG in relation to OTG. The number of chondrocytes did not show any significant difference. The nuclear volume of chondrocytes in the superficial layer was higher in CG than in OSG . In the medium layer, the volume of chondrocytes was higher in CG than in OSG and the OTG. The same occurred when the values of the epiphysial cartilage was considered as a whole. For the collagen, the values of volume density of the collagen fibers were higher in the OSG and in the OTG than the CG. The same occurred with the OTG in relation to OSG. Conclusions The depression of estrogen levels produced important changes in the tibial cartilage of Wistar rats mainly on the superficial layer and physical activity seemed to relieve these effects. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT PURPOSE Pellucid marginal degeneration (PMD) of the cornea is a rare ectatic disorder which typically affects the inferior or superior peripheral cornea in a crescentic fashion. We report histological and ultrastructural features of three PMD corneas. METHODS: The following three patients were diagnosed with PMD corneas: (A) one 41-year-old male, (B) one 56-year-old female, and (C) one 31-year-old male. The patients underwent keratoplasty and the excised corneas were processed for light and electron microscopy to study the ultrastructural features. RESULTS: Degenerated corneas were observed in the region adjacent to the limbus. In the degenerated region of the cornea, the Bowman's layer had been replaced by collagenous pannus and the anterior stroma contained degenerated collagen fibrils (CFs) with very large proteoglycans (462±420 nm 2 ). The lamellae were fused and keratocytes appeared like fibroblast. The prelimbal region of the PMD cornea had a degenerated Bowman's layer and thin undulating lamellae in the stroma. The CFs of the Bowman's layer and the stroma were replaced by very fine microfilaments. The mean of the minimum CF diameter was 19±3 nm in London Rsin White-embedded tissue. CONCLUSIONS: Our observations of the disorganization and degeneration of CFs suggest that PMD could be related to a disorder in the synthesis of CF. This disorder was more severe in the cornea adjacent to the limbus compared to the cornea further away from the limbus. Microsc. Res. Tech., 2013. © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT Colon cancer classification has a significant guidance value in clinical diagnoses and medical prognoses. The classification of colon cancers with high accuracy is the premise of efficient treatment. Our task is to build a system for colon cancer detection and classification based on slide histopathological images. Some former researches focus on single label classification. Through analyzing large amount of colon cancer images, we found that one image may contain cancer regions of multiple types. Therefore, we reformulated the task as multi-label problem. Four kinds of features (Color Histogram, Gray-Level Co-occurrence Matrix, Histogram of Oriented Gradients and Euler number) were introduced to compose our discriminative feature set, extracted from our dataset that includes six single categories and four multi-label categories. In order to evaluate the performance and make comparison with our multi-label model, three commonly used multi-classification methods were designed in our experiment including one-against-all SVM (OAA), one-against-one SVM (OAO) and multi-structure SVM. Four indicators (Precision, Recall, F-measure, and Accuracy) under 3-fold cross-validation were used to validate the performance of our approach. Experiment results show that the precision, recall and F-measure of multi-label method as 73.7%, 68.2%, and 70.8% with all features, which are higher than the other three classifiers. These results demonstrate the effectiveness and efficiency of our method on colon histopathological images analysis. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT The palatine mucosa and filiform papillae of the dorsal tongue mucosae of rodents were examined using transmission electron microscopy (TEM) and high resolution scanning electron microscopy (HRSEM). In the HRSEM method, the samples were fixed in 2% osmium tetroxide, dehydrated in alcohol, critical point-dried, and coated with gold-palladium. In addition, the HRSEM technique was used for morphometric analysis (length, width, and length/width ratio of cocci and bacilli). For the TEM method, the tissues were fixed in modified Karnovsky solution (2.5% glutaraldehyde, 2% formalin in 0.1 M sodium phosphate buffer, pH 7.4) and embedded in Spurr resin. The results demonstrated that there are thick polygonal keratinized epithelial cells where groups of bacteria are revealed in three-dimensional images on the surface of filiform papillae in these animals. The bacterial membranes are randomly attached to the microplicae surface of epithelial cells. Morphometrics showed higher values of length and width of cocci in newborn (0 day) as compared to newborn (7 days) and adults animals, the bacilli showed no differences in these measurements. At high magnification, the TEM images revealed the presence of glycocalyx microfilaments that constitute a fine adhesion area between bacterial membranes and the membranes of epithelial microplicae cells. In conclusion, the present data revealed the fine fibrillar structures of bacteria that facilitate adhesion to the epithelial cell membranes of the oral cavity and morphometric changes in newborn (0 day) rats as compared with other periods. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT We propose a light sheet based imaging flow cytometry technique for simultaneous counting and imaging of cells on a microfluidic platform. Light sheet covers the entire microfluidic channel and thus omits the necessity of flow focusing and point scanning based technology. Another advantage lies in the orthogonal detection geometry that totally cuts-off the incident light, thereby substantially reducing the background in the detection. Compared to the existing state-of-art techniques the proposed technique shows marked improvement. Using fluorescently-coated Saccharomyces cerevisiae cells we have recorded cell counting with throughput as high as 2,090 cells/min in the low flow rate regime and were able to image the individual cells on-the-go. Overall, the proposed system is cost-effective and simple in channel geometry with the advantage of efficient counting in operational regime of low laminar flow. This technique may advance the emerging field of microfluidic based cytometry for applications in nanomedicine and point of care diagnostics. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT Based on its histochemical properties, the secretory portion of the hamster submandibular gland has been classified as seromucous cells. The presence of endogenous peroxidase (PO) reaction was shown in the nuclear envelope, cisternae of endoplasmic reticulum and Golgi apparatus. The 3,3′-diaminobenzidene, tetrahydrochloride (DAB) method revealed bipartite secretory granules containing a PO-positive dense core surrounded by a less dense halo in these cells. In the present investigation, serous and mucous-like cells were found in resin-embedded semi-thin sections of the DAB-reacted hamster submandibular gland. These sections were already on glass slides for routine light microscopic observations, therefore electron microscopic analysis could be unrealizable. We then used reflectance-mode confocal laser scanning microscopy to visualize additional sites of PO activity as detected in these sections. Using this approach, we found mucous cells with PO activity-negative secretory granules and seromucous cells with PO activity-positive spot-like secretory granules of the regular sublingual gland most frequently adjacent to the serous cells with typical electron-dense secretory granules. These cells clearly differ from the seromucous cells with bipartite secretory granules and the granular duct cells with typical electron-dense secretory granules of the hamster submandibular gland. Additionally, secretory endpieces of the ectopic sublingual gland-like tissue empty into the duct of the hamster submandibular gland lobule. Thus, our findings suggest that a mass of sublingual gland tissue extends into the hamster submandibular gland during its development, and PO may be synthesized and secreted into the same duct. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT The aim of this study was to evaluate the biocompatibility in rat subcutaneous connective tissue of a new zinc oxide endodontic sealer (Endomethasone N) compared to those provided by Endofill and Sealer 26. Polyethylene tubes containing the test materials were implanted into dorsal subcutaneous connective tissue of Wistar albino rats. After 7 and 42 days, the implants with the surrounding tissue were collected, fixed, and processed for histologic evaluation. Sections were evaluated for the presence of inflammatory cells (poly or monomorfonuclear), blood vessels, necrosis area, and thickness of fibrous capsule. Comparisons between groups and time-periods were performed with Kruskal–Wallis and Mann–Whitney U non-parametric tests for 5% significance level. No differences in the biocompatibility patterns among the materials for the 2 experimental periods were observed. Independently of the sealer, the tissue behavior showed a tendency to decrease the irritation effect over time. It can be concluded that all sealers are irritant, but its toxicity decreased with time. Endomethásone N showed biocompatible characteristics comparable with those provided by Endofill and Sealer 26. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: We report on the microstructure and interfacial chemistry of thin films of pure and La-doped multiferroic bismuth ferrite (Bi 1-x La x FeO 3 or BLFO), synthesized on Indium Tin Oxide-coated glass substrates by solution-deposition technique and studied using scanning transmission electron microscopy. Our results show that undoped and La-doped thin films are polycrystalline with distorted rhombohedral structure without any presence of any line or planar defect in the films. In addition, the films with La doping did not show any structural change and maintain the equilibrium structure. Cross section compositional analysis using X-ray energy dispersive spectrometry did not reveal either any interdiffusion of chemical species or formation of reaction product at the film-substrate interface. However, a closer examination of the microstructure of the films shows tiny pores along with the presence of ∼2–3 nm thin amorphous layers, which may have significant influence on the functional properties of such films. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT Two-photon microscopy is an innovative technology that has high potential to combine the examination of soft and hard tissues in vitro and in vivo . Calcium phosphates are widely used substitutes for bone tissue engineering, since they are degradable and consequently replaced by newly formed tissue. It is well known that osteoclasts are responsible for the resorption processes during bone remodelling. We hypothesize that also macrophages are actively involved in the resorption process of calcium phosphate scaffolds and addressed this question in in vitro culture systems by two-photon laser scanning microscopy. Beta-tricalcium phosphate specimens were incubated with (1) macrophages, (2) interleukin-4 activated macrophages, and (3) osteoclasts for up to 21 days. Interestingly, macrophages degraded beta-tricalcium phosphate specimens in an equivalent fashion compared to osteoclasts and significantly more than IL-4 activated macrophages. An average of ∼32% of the macrophages was partially filled with ceramic material while this was 18% for osteoclasts and 9% for IL-4 activated macrophages. For the first time by applying two-photon microscopy, our studies show the previously unrecognized potential of macrophages to phagocytose ceramic material, which is expected to have implication on osteoconductive scaffold design. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT Hastelloy N alloy has been selected as the primary structure material for molten salt reactor. In this article, Hastelloy N alloy samples were irradiated to different doses at room temperature using 300 keV Ar + ions. The microstructural evolution was investigated by transmission electron microscopy (TEM) and energy-dispersive spectroscopy (EDS). Black dot defects emerged in sample irradiated at low dose (0.4 displacement per atom (dpa)), and they grew up with irradiation doses (0.4–2 dpa). A high density of small dislocation loops (nano meters in size) were observed in the sample irradiated to 4 dpa. When the ion dose increased to 12 dpa, complicated structures with defects (including dislocation lines, larger loops and smaller black dots) were observed. Dislocation networks were detected from high-angle annular dark field (HAADF) images. Larger dislocation loops (size: 30–80 nm) were visible in the sample irradiated to 40 dpa. Irradiation with dose of 120 dpa led to the formation of face-centered cubic nanocrystallites with preferred orientations. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT Scanning electron microscopy on the postembryonic development of the mouthpart sensory structures of the Muga silk moth, Antheraea assamensis revealed the presence of a variety of sensilla. The types, distribution pattern of the sensilla remained almost the same in the different larval stages except for the number and dimension in some cases. The findings of the study assumes significance because the silk moth, Antheraea assamensis is a species endemic to the North Eastern part of India and very few reports are available on it. In fact, the present study provides the first report on the ontogeny of the mouth part sensilla of the silk moth species. Since mouth part sensilla and their surface micro structural characteristics are extremely important in understanding their roles in searching behavior during feeding, the present study will certainly help in developing strategies for proper rearing of this economically important insect species. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT Novel monodisperse mesoporous iron oxide nanoparticles (m-IONPs) were synthesized by a postsynthesis etching approach and characterized by electron microscopy. In this approach, solid iron oxide nanoparticles (s-IONPs) were first prepared following a solvothermal method, and then etched anisotropically by polyacrylic acid to form the mesoporous nanostructures. MTT cytotoxicity assay demonstrated that the m-IONPs have good biocompatibility with mesenchymal stem cells (MSCs). Owing to their mesoporous structure and good biocompatibility, these monodisperse m-IONPs were used as a nonviral vector for the delivery of a gene of vascular endothelial growth factor (VEGF) tagged with a green fluorescence protein (GFP) into the hard-to-transfect stem cells. Successful gene delivery and transfection were verified by detecting the GFP fluorescence from MSCs using fluorescence microscopy. Our results illustrated that the m-IONPs synthesized in this work can serve as a potential nonviral carrier in gene therapy where stem cells should be first transfected and then implanted into disease sites for disease treatment. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT The protein organization within focal adhesions has been studied by state-of-the-art super resolution methods because of its thin structure, well below diffraction limit. However, to achieve high axial resolution, most of the current approaches rely on either sophisticated optics or diligent sample preparation, limiting their application. In this report we present a phasor-based method that can be applied to fluorescent samples to determine the precise axial position of proteins using a conventional confocal microscope. We demonstrate that with about 4,000 photon counts collected along a z-scan, axial localization precision close to 10 nm is achievable. We show that, with within 10 nm, the axial location of paxillin, FAK, and talin is similar at focal adhesion sites, while F-actin shows a sharp increase in height towards the cell center. We further demonstrated the live imaging capability of this method. With the advantage of simple data acquisition and no special instrument requirement, this approach could have wide dissemination and application potentials. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT The aim of this study was to evaluate the effect of final irrigation protocols (17% EDTA, BioPure MTAD, SmearClear, and QMiX) on microhardness and erosion of root canal dentin. Fifty roots were sectioned transversely at the cement–enamel junction and each root was sectioned horizontally into 4-mm-thick slices. The samples were divided into five groups ( n  = 10) according to the final irrigation protocol: G1: distilled water (control group); G2: 17% EDTA; G3: BioPure MTAD; G4: SmearClear; and G5: QMiX. The dentin microhardness was then measured with a load of 25 g for 10 s. Initially, the reference microhardness values were obtained for the samples without any etching. The same samples were then submitted to the final irrigation protocols. A new measure was realized and the difference between before and after the procedures was the dentin microhardness reduction. In sequence, the specimens were submitted to SEM analysis to verify the dentinal erosion. The Kruskal Wallis and Dunn tests ( α  = 5%) were used to compare the results. The dentin microhardness decreased for all final irrigation protocols. There was no significant difference between groups 2, 3, 4, and 5 ( P  〉 0.05), but this groups presented significant dentin microhardness reduction than G1 ( P  〈 0.05). In G2, occurred the highest incidence of dentinal erosion ( P  〈 0.05). 17% EDTA, BioPure MTAD, SmearClear, and QMiX promoted significant dentin microhardness reduction. Dentinal tubules erosion was promoted by 17% EDTA. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT The aim of this study was to investigate the effect of Sibutramine on platelet ultrastructure and discuss the morphological observations in relation to known physiological effects of the compound. Six-week-old, female Spraque-Dawley rats were used in this study. The animals were placed on a high energy diet after which sibutramine administration followed. Blood was drawn on the day of termination and platelet rich plasma was obtained to prepare plasma smears for analysis. Scanning electron microscopy was used to investigate the ultrastructure of the platelets. Platelets of the Sibutramine-treated animals showed smooth surface with limited pseudopodia formation when compared with that of the control animals. Higher magnification of the platelet surface showed membrane tears and swelling, typically seen in necrotic cells. It can therefore be concluded from these results that Sibutramine alters the membrane morphology of platelets to that typical of necrotic cells. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT Retroviral transduction of four transcription factors (Oct4, Sox2, Klf4 and c-Myc) or three factors, excluding c-Myc, has been shown to initiate a reprogramming process that results in the transformation of murine fibroblasts to induced pluripotent stem (iPS) cells, and there has been a rapid increase in the number of iPS cell-based preclinical trials. In this study, the effects of these transcription factors were evaluated regarding the growth and differentiation of murine iPS cells under hypoxia. Based on the results of RT-PCR and alizarin red S staining, there were no statistical differences in the growth and differentiation of iPS cells or the induction of iPS cells to osteoblasts under hypoxia between the transcription factor groups. Furthermore, the function of hypoxia inducible factors (HIFs) in murine iPS cells under hypoxia was investigated in relation to the morphology and expression of transcription factors using RT-PCR and Western blotting. The HIF-2α knockdown group exhibited a decrease in the colony size of the iPS cells. The HIF-2α or -3α knockdown group demonstrated a statistically significant decrease in the transcription factor expression compared to that observed in the control group. These results demonstrate that HIF-2α among HIFs is the most influential candidate for the maintenance of the pluripotency of murine iPS cells. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT Fixation of biological samples is an important process especially related to histological and ultrastructural studies. Chemical fixation was the primary method of fixing tissue for transmission electron microscopy for many years, as it provides adequate preservation of the morphology of cells and organelles. High pressure freezing (HPF) and freeze substitution (FS) is a newer alternative method that rapidly freezes non-cryoprotected samples that are then slowly heated in the FS medium, allowing penetration of the tissue to insure adequate fixation. This study addresses several issues related to tissue preservation for electron microscopy. Using mice liver tissue as model the difference between samples fixed chemically or with HPF immediately after excision, or stored before chemical or HPF fixation were tested with specific focus on the nuclear membrane. Findings are that immediate HPF is the method of choice compared to chemical fixation. Of the chemical fixatives, immediate fixation with 2.5% glutaraldehyde (GA)/formaldehyde (FA) is the best in preserving membrane morphology, 2.5% GA can be used as alternative for stored and then chemically processed samples, with 10% formalin being suitable as a storage medium only if followed by HPF fixation. Overall, storage leads to lower ultrastructural preservation, but HPF with FS can minimize these artifacts relative to other processing protocols. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT Immunopresence of estrogen receptor α (ERα), β (ERβ) and progesterone receptor (PR) were examined in the ewe mammary gland from prepubertal stage to involution. Immunolocalization of ERα revealed a strong positive staining in nuclei of cells composing terminal ductular units (TDUs) in prepubertal ewes. A mild immunoreactivity was identified in early lactating gland. During late lactation immunoreactive product to ERα was observed in the cytoplasm of glandular epithelial cells in all alveoli. In mammary glands at involution ERα positivity was clearly nuclear, with weak to moderate cytoplasmic staining. Cytoplasmic strong immunostaining for ERβ was detected in cells of TDUs, whereas some stromal cells exhibited nuclear staining. A nuclear ERβ immunostaining was observed at early lactation, instead during late lactation, the positivity for ERβ showed only a moderate cytoplasmic distribution. At involution, ERβ positivity was very moderate and detected just in the cytoplasm of shrunken alveoli. Scattered nuclear staining of PR was observed just in mammary glands at early lactation. These results showed that in the mammary glands of sheep both estrogen receptor isoforms were displayed during lactation cycle and that PR appeared just at early lactation, reflecting their regulatory role in alveolar cells. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT Generally, shape from focus methods use a single focus measure to compute focus quality and to obtain an initial depth map of an object. However, different focus measures perform differently in diverse conditions. Therefore, it is hard to get accurate 3D shape based on a single focus measure. In this article, we propose a total variation based method for recovering 3D shape of an object by combining multiple depth hypothesis obtained through different focus measures. Improved performance of the proposed method is evaluated by conducting several experiments using images of synthetic and real microscopic objects. Comparative analysis demonstrates the effectiveness of the proposed approach. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT The roots of Salvia miltiorrhiza Bge., named Danshen in traditional Chinese medicine, is widely used for the treatment of cardiovascular disorders, also for food auxiliary agent. Rosmarinic acid, salvianolic acid B, dihydrotanshinone, cryptotanshinone, and tanshinone IIA are all responsible for the pharmacological activity of Danshen, they are important quality evaluation index for Danshen. The quality evaluation of Danshen is mainly concentrated in difference of cultivated regions and years, but the difference in root systems themselves and the cause of the difference were overlook. The growth rings were employed to identified the roots ages in root systems. Then the contents of eight bioactive component in different aged, diameter and tissue of individual roots of the same root system were evaluated using HPLC coupled with an ultraviolet photodiode array detector. The root system of S. miltiorrhiza was a complex root systems composed by different aged and diameter roots, in which the contents of bioactive compounds vary greatly from the other. The lipophilic diterpenoid quinones mainly distributed in the bark and increased with the cultivated years. The hydrophilic depsides distributed in all tissues, and the contents are relatively high in xylem in 1-year-old roots. It was very important to identify the individual root age of S. miltiorrhiza root system cultivated for several years. In the same root system of S. miltiorrhiza roots with 0.4-cm diameter contain more bioactive compounds, which maybe cause by the uneven distribution of bioactive compounds. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT Eucryptorrhynchus chinensis (Olivier) (Coleoptera: Curculionidae) and E. brandti (Harold) (Coleoptera: Curculionidae) are the two most important pests of tree-of-heaven, Ailanthus altissima (Mill.) Swingle and its variety Ailanthus altissima var. Qiantouchun in China. They are also considered potential biological control agents for tree-of-heaven in North America. In this study, the external morphologies and antennal sensilla of both species were examined using scanning electron microscopy to better understand their host-finding mechanisms. Eleven morphological sensilla types were recorded, that is, Böhm bristles, six types of sensilla chaetica (Sch. 1–6), two types of sensilla basiconica (Sb. 1–2), and two types of sensilla trichodea (St. 1–2). Sch. 5 were absent from the antennae of E. chinensis , while Sch. 2 were absent from the antennae of E. brandti . Abundant cuticular pores were present on the antennae of both species. Three types of sensilla on the antennae of E. chinensis that were not found in a previous study, and ten different types of sensilla on the antennae of E. brandti were identified for the first time. The possible functions of the sensilla types are discussed based on a comparison with previous studies. Four types of sensilla (Sb. 1, Sb. 2, St. 2, and Sch. 6) on the antennae of both species indicate chemoreception may play a significant role in host location. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT Myotis nigricans is an endemic species of vespertilionid bat, from the Neotropical region, that resembles temperate zone bats in their reproductive cycle; presenting an annual reproductive cycle with two periods of testicular regression, which are not linked to the apoptotic process and seems to be not directly linked to any seasonal abiotic variation. Thus, this study aimed to ultrastructurally evaluate their reproductive cycle. The process of testicular regression could be divided into four periods: active; regressing; regressed and recrudescence; with all presenting distinct characteristics. The active period was similar to that of other bats, presenting the complete occurrence of spermatogenesis, with three main types of spermatogonia (A d , A p , and B) and 12 steps in spermatid differentiation; however, it differed in having the outer dense fibers 1, 5, 6, and 9 larger than the others. These three types of spermatogonia undergo considerable morphologic changes from regressing to the regressed period, and in the recrudescence, they return to the basic morphology, which reactivates spermatogenesis. In conclusion, our study described the process of spermatogenesis, the ultrastructure of the spermatozoa and the distinct morphologic variations in the ultrastructure of the testicular cells of M. nigricans during the four different periods of its annual reproductive cycle. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT The extraction efficiency of evanescent light from ZnO nanolayers and their thickness profiles in the range of (1–105) nm was evaluated by a new microscopy technique, differential evanescent light intensity imaging method. It is based on capturing the evanescent light scattered by the layer of the material deposited on glass substrates. The analyzed ZnO films were obtained by pulsed laser deposition at 27°C and 100°C, using a nanosecond UV laser source. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT Microorganisms are able to survive and induce persistent infection in extraradicular areas. The objective of this study was to evaluate the relationship between extraradicular biofilm and persistent periapical periodontitis. Thirty-five apical samples with different stages of pulp and periapical pathology (vital pulp, pulp necrosis without radiographically visible periapical lesions, chronic periapical periodontitis that had not received root canal therapy and persistent periapical periodontitis) were initially evaluated using scanning electron microscopy. The same samples were then processed using Brown and Brenn-modified Gram staining. We detected extraradicular biofilm in all samples with persistent periapical periodontitis and in three samples with chronic periapical periodontitis. The extraradicular bacteria predominantly had rod and filament morphology and were surrounded by varying amounts of amorphous extracellular material. The surfaces outside the root of the apical samples with vital pulp and pulp necrosis were covered by fibers, and no extraradicular microorganisms were present, which suggests that extraradicular biofilm is closely associated with failed endodontic treatments, thus resulting in persistent infection. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT Aspects of ionoregulatory or mitochondria-rich cell (MRC) differentiation and adaptation in Nile tilapia yolk-sac larvae following transfer from freshwater to elevated salinities, that is, 12.5 and 20 ppt are described. Investigations using immunohistochemistry on whole-mount Nile tilapia larvae using anti- Na + /K + -ATPase as a primary antibody and Fluoronanogold™ (Nanoprobes) as a secondary immunoprobe allowed fluorescent labeling with the high resolution of confocal scanning laser microscopy combined with the detection of immunolabeled target molecules at an ultrastructural level using transmission electron microscopy (TEM). It reports, for the first time, various developmental stages of MRCs within the epithelial layer of the tail of yolk-sac larvae, corresponding to immature, developing, and mature MRCs, identifiable by their own characteristic ultrastructure and form. Following transfer to hyperosmotic salinities the density of immunogold particles and well as the intricacy of the tubular system appeared to increase. In addition, complementary confocal scanning laser microscopy allowed identification of immunopositive ramifying extensions that appeared to emanate from the basolateral portion of the cell that appeared to be correlated with the localization of subsurface tubular areas displaying immunogold labeled Na + /K + -ATPase. This integrated approach describes a reliable and repeatable prefixation immunogold labeling technique allowing precise visualization of NaK within target cells combined with a 3D imaging that offers valuable insights into MRC dynamics at an ultrastructural level. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT The morphology of the trigeminal ganglion in human fetus was investigated by means of the tract-tracing method using the lipophilic dye DiI-C18-(3) (1,1′-double octadecane 3,3,3′3′-tetramethyl indole carbonyl cyanine-perchlorate), hematoxylin–eosin (HE) stain, and three-dimensional computer reconstruction models. The trigeminal ganglion was flat in the dorsoventral direction, and DiI staining revealed that the trigeminal ganglion cells were somatotopically distributed in the ganglion in a way that reflected the mediolateral order of the three branches. Ganglion cells of the ophthalmic nerve were distributed in the anteromedial part of the trigeminal ganglion, those of the mandibular nerve were in the posterolateral part, and those of the maxillary nerve were localized in the intermediate part. DiI labeled both ganglion cells and nerve fibers in the trigeminal ganglion; the ganglion cells varied in size and appeared as round- or oval-shaped, the neurites connected the cell soma, and some bipolar neurons were also observed. The number of embryonic trigeminal ganglion cells did not significantly change with gestational age, but the cell diameter, area, and perimeter significantly increased. The motor root leaves the pons, runs along the sensory root, passes the ventral surface of the ganglion, and finally runs together with the mandibular nerve. The findings reported here elucidate the morphology, development, and somatotopic organization of the trigeminal ganglion and reveal the trigeminal nerve motor root pathway along the trigeminal ganglion and mandibular nerve in the human fetus. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: A cross-section sample preparation technique is described for transmission electron microscopy studies of metallic materials. The technique uses jet electro-polishing for the final perforation. Examples are provided of using this technique for copper-support/copper-films/copper-support multilayer structures, grown by electro-deposition. The samples prepared by our current technique are compared with the ones made by ion-milling. The technique is also applicable to materials which are susceptible to ion beam and thermal damages. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT Haematoxylin/Eosin staining was performed to screen for Hirano bodies from the temporal lobe including the hippocampus in 10 Pick's disease cases containing Pick bodies. Although the inclusions were confirmed in 9 out of 10 cases, only 4 out of 10 were particularly enriched with the eosinophilic bodies. These were subjected to immunostaining with anticomplement antibodies and astrocyte marker antiglial fibrillary acidic protein antibody and the HLA class II CR3/43 antibody to visualize microglia. An intraneuronal Hirano body was observed in one case that otherwise contained mainly the extracellular inclusions. In all cases, the extracellular Hirano bodies were seen lying adjacent to soma of neurons within CA1 region of the hippocampus. The extracellular Hirano bodies stained intensely with C1q, the first component of the classical pathway of activation but remained unreactive against antibodies to C4 and the C3 activation products (C3b and iC3b) and the alternative complement pathway component factor B. Hirano bodies also remained negative with the antiglial fibrillary acidic protein for astrocytes and HLA class II antibody CR3/43 for microglia. The results demonstrate that Hirano bodies have strong immunoreactivity to C1q; however, whether other complement components are associated with these inclusions remains to be further investigated. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT This study evaluated the synergy of professional acidulated fluoride gel (APF) or fluoridated dentifrice application combined with Er,Cr:YSGG laser irradiation on the formation of CaF 2 -like products (CaF 2 ), in vitro. Thus, 272 bovine enamel slabs were randomly distributed among eight groups: G1: untreated enamel; G2: treated with fluoridated dentifrice (NaF, 1,100 μgF/g); G3: treated with acidulated phosphate fluoride gel (APF, 1.23% F - ); G4: irradiated with Er,Cr:YSGG laser at 8.5 J/cm 2 ; G5 and G6: combination of pre-irradiation with Er,Cr:YSGG followed by dentifrice or APF application, respectively; G7: combination of dentifrice application followed by Er,Cr:YSGG irradiation; G8: combination of APF application followed by Er,Cr:YSGG irradiation. After treatments, samples were evaluated by scanning electron microscopy, and the content of CaF 2 was determined by an ion specific electrode. Both APF and dentifrice application promoted the formation of CaF 2 on enamel, whereas Er,Cr:YSGG irradiation promoted an increase of roughness of the enamel, increasing the surface area. Laser irradiation before fluoridated products increased the content of CaF 2 formed when compared to groups that APF or dentifrice were applied isolated. However, the content of CaF 2 formed when irradiation was performed after APF or dentifrice was not statically significant when compared to the control groups. In conclusion, Er,Cr:YSGG laser increases the formation of CaF 2 on enamel when the irradiation is performed before the application of APF or dentifrice. The association of laser with APF is most promissory for caries prevention because of the higher concentration of CaF 2 formation and also the chemical changes promoted by laser irradiation demonstrated in literature. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT We report a quick and easy method for a random selected area electron diffraction (SAED) pattern without rotating and tilting the specimen to perform phase identification and unit cell determination by combined with the XRD softwares. If your TEM is well aligned and camera length is carefully corrected, two-dimensional (2D)-SAED pattern can be directly transformed to 1D-profile after the center determination of pattern, this profile is then imported to XRD analysis packages. Finally, phase identification and unit cell determination can be performed after peak search or precise peak position determined by profile fitting. Two examples, flaky-like TiO 2 nanomaterial and TiO 2 nanotubes precipitated by the silver nanoparticles, were tested and verified for the validation of phase identification and unit cell determination using this method; the successful crystallographic analysis of one single gold nanocrystal indicates it is still validate for the nanocrystals with the smaller diffraction volume, but need two or more random tilt SAED patterns. This method could be further used in the quantitative phase analysis, structure determination and Rietveld refinement for the nanomaterials if the reliable integrated intensity can be extracted. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT Diagnostic hysteroscopy is a popular method for investigating the regions in the female reproductive system. The videos generated by hysteroscopy sessions of patients are recurrently archived in medical libraries. Gynecologists often need to browse these libraries in search of similar cases or for reviewing old videos of a patient. Diagnostic hysteroscopy videos contain a lot of information with abundant redundancy. Key frame extraction-based video summarization can be used to reduce this huge amount of data. Moreover, key frames can be used for browsing and indexing of hysteroscopy videos. In this article, a domain specific visual attention driven framework for summarization of hysteroscopy videos is proposed. The visual attention model is materialized by computing saliency based on color, texture, and motion. The experimental results, in comparison with other techniques, demonstrate the efficacy of the proposed framework. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT The article studies thermal resistance variation by analyzing the colorimetric parameters correlated with the optical microscopy data of two groups of ceramic pigments obtained by co-precipitation in aqueous medium of phosphate anion and of a mixture of chromium phosphate with various chromophore cations. This research enabled us to reveal the thermal thresholds/domains within which significant color changes occur, thus allowing the choice of pigments compatible with the thermal varnishing-glazing processes involved in the manufacture of tesserae for mosaic and stained glass and of colored materials for floor tiles, wall tiles and painted porcelain. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT The use of cyclosporine (CsA) has shown to induce an increase in density of oligodendrocytes near remyelinating areas following the injection of ethidium bromide (EB), a demyelinating agent, in the rat brainstem. It is also known that diabetes mellitus was capable of delaying remyelination by both oligodendrocytes and Schwann cells in this gliotoxic model. This study was designed to assess whether CsA had the capacity to improve remyelination in streptozotocin-induced (50 mg/kg, intraperitoneal route) diabetic rats. Diabetic Wistar rats were divided in different groups receiving 10 microlitres of 0.1% EB or 0.9% saline solution into the cisterna pontis and were treated or not with CsA. During 7 days and, thereafter, three times a week, 10 mg/kg/day of CsA were given by intraperitoneal route. The rats were euthanized from 7 to 31 days after EB or saline injection and brainstem sections were collected and processed for light and transmission electron microscopy studies. Results from different groups were compared by using a semi-quantitative method developed for documenting the extent and nature of remyelination in semithin sections following gliotoxic lesions. Results showed that CsA administration to diabetic rats after EB injection stimulate both oligodendroglial and Schwann cell remyelination (mean remyelination scores of 3.15 ± 0.5 for oligodendrocytes and 1.36 ± 0.58 for Schwann cells) compared to untreated animals (2.52 ± 0.71 for oligodendrocytes and 0.73 ± 0.47 for Schwann cells, respectively). CsA given to diabetic rats was capable of reversing some of the deleterial effects of diabetes on remyelination. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT  Background : Polymethylmethacrylate used in surgery is one of the first biomaterials. Conventional histology dissolves the resin; one of the reasons that only few complete histology is published. Objectives and Purpose : The question is, whether a complete histology changes the understanding, influences the application and opens approaches for improvements. The dos and don'ts of the processing technology are presented in reproducible manner. Material and Methods : Ten femurs of giant rabbits were taken from running experiments of femur-canal filling with bone cement. Different stages were considered for high-resolution histology and electron microscopy: 4-weeks-stage (bone healing), 12-weeks-stage (remodelling) and one and two years. A human-cadaver specimen with a follow up of two years was processed. All animals were perfusion-fixated and the complete vasculature micro-casted. Serial cuts were performed with a stone saw, followed by a wet grinding processing. The fluorescence documentation in the High Intensity Incident Fluorescent Light (HIIFL) and Orthoplan Leitz Ploemopak ® was applied and high resolution microradiography used the Siemens Kristalloflex ® . The Scanning Electron Microscopy (SEM) was performed applying deep-freezing technology for the PSEM-500. The human specimen was embedded after sectioning using epoxy-resin. Results : All cement implants showed osseointegration and remodelling with a tangential adherence of bone onto the ceramic/PMMA surfaces. Intact cancellous structures after one and two years did not show any signs of heat necrosis. The human specimen confirmed the results from the animal experiments. Conclusion : The complete bone-to-cement histology changed the understanding of the bone cements function, influenced its application and opened new ways for improvement. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT Actually, the most used method to control ticks is synthetic acaricides with neurotoxic action. However, the use of these methods presents inconveniences, such as the contamination of the environment and risks to the host's health due to the residual effects. Thus, several studies have been developed aiming to find alternative ways to control these ectoparasites, such as the use of natural compounds with active ingredients, which act controlling some species of plagues in addition to presenting medicinal properties that are beneficial to humans. The present study aimed to analyze the action of andiroba oil ( Carapa guianensis ) on the synganglion of Rhipicephalus sanguineus semiengorged females through morphological and cytochemical techniques aiming to verify if this natural product have neurotoxic action as the numerous synthetic acaricides. The results showed that andiroba oil interferes in the synganglion through structural and enzymatic changes, which lead the nervous tissue to apoptotic death involving autophagy. Among these changes was observed the emergence of large empty spaces between the perineurium and the cortical region, vacuolated cortex cells and with cell swelling, neural cells with picnotic nuclei or in initial stage of chromatin margination and neuropile with high structural disorganization. Considering these data, it can be concluded that andiroba seed oil can be used as an alternative method in the control of R. sanguineus ticks due to its neurotoxic action. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT Transparent conducting Mn-doped ZnO thin films have been prepared by successive ionic layer by adsorption reaction (SILAR) method. The deposition conditions have been optimized based on their structure and on the formation of smoothness, adherence, and stoichiometry. The results of the studies by X-ray diffraction, scanning electron microscope (SEM), reveal the varieties of structural and morphological modifications feasible with SILAR method. The X-ray diffraction patterns confirm that the ZnO:Mn has wurtzite structure. The interesting morphological variations with dopant concentration are observed and discussed. The films' quality is comparable with those grown with physical methods and is suitable for spintronic applications. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT Conducting atomic force microscopy and scanning surface potential microscopy were used to study the local electrical properties of gallium-doped zinc oxide (GZO) films prepared by pulsed laser deposition (PLD) on a polyimide (PI) substrate. For a PLD deposition process time of 8 min, the root-mean-square roughness, coverage percentage of the conducting regions, and mean work function on the GZO surface were 2.33 nm, 96.6%, and 4.82 eV, respectively. When the GZO/PI substrate was used for a polymer light-emitting diode (PLED), the electroluminescence intensity increased by nearly 20% compared to a standard PLED, which was based on a commercial-ITO/glass substrate. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT Daqingye and Banlangen are commonly used Chinese medicinal materials derived from the leaves and roots of Isatis indigotica Fort., respectively, which clinical effects have been confirmed by many studies in recent years. However, many problems have arisen concerning the quality and identity of materials sold in the market under these two names. Thus, the identification of Daqingye and Banlangen has drawn public attention. In this work, transverse sections of Daqingye and Banlangen from I . indigotica Fort. and two easily confused species, namely Baphicacanthus cusia (Nees) Bremek. and Clerodendrum cyrtophyllum Turcz., were investigated with normal light and fluorescence microscopy. The distinguishing features were 7–9 vascular bundles, cystoliths and nonglandular hairs in the leaves of I . indigotica , B . cusia , and C . cyrtophyllum , respectively. The Banlangen could be distinguished according to the characteristics of parenchymous cells, cystoliths, and stone cells. Moreover, the fluorescence features of Daqingye and Banlangen investigated in this study can provide direct points for differentiating those samples. Importantly, whether the crude drugs were decocted could be easily identified by their different fluorescence features, which can ensure their quality in clinical application. This is the first report to distinguish the three species that are commonly found in the market sold as Daqingye and Banlangen by normal light and fluorescence microscopy. This work indicates that the combination of normal light and fluorescence microscopy could be powerful, convenient, and economical for authenticating Daqingye and Banlangen from the three species, including crude drugs and decoction dregs. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT Atomic force microscopy (AFM) proved to be able to obtain high-resolution three-dimensional images of single-membrane proteins, isolated, crystallized, or included in reconstructed model membranes. The extension of this technique to native systems, such as the protein immersed in a cell membrane, needs a careful manipulation of the biological sample to meet the experimental constraints for high-resolution AFM imaging. In this article, a general protocol for sample preparation is presented, based on the mechanical stretch of the cell membrane. The effectiveness for AFM imaging has been tested on the basis of an integrated optical and AFM approach and the proposed method has been applied to cells expressing cystic fibrosis transmembrane conductance regulator, a channel involved in cystic fibrosis, showing the possibility to identify and analyze single proteins in the plasma membrane. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT It is well known that estrogen is neuroprotective through various mechanisms which suggest that sex hormone levels, thrombotic mechanisms, and inflammatory processes are strongly interconnected in predicting the outcome and consequences of cerebral ischemia. Because platelet ultrastructure is altered in conditions like thrombosis and associated with stroke, the question arises whether ultrastructural analyses of platelet morphology may provide further insight into the role of estrogen during ischemic insult. In the current study, a hyperglycemic modification to the two-vessel occlusion model for inducing experimental cerebral ischemia was employed, in order to correlate neural tissue integrity levels between three experimental groups to corresponding platelet ultrastructure so as to determine whether there is an association between cerebral ischemia and the presence of inflammatory or necrotic platelet ultrastructure. It is apparent in the results that under the influence of estrogen in cyclic or intact females, there is lesser neural tissue damage as well as a reduced degree of inflammation evident in platelet activation morphology when compared to males and acyclic or ovariectomized females. It is unmistakable that neural injury is closely shadowed, if not preceded, by inflammatory changes in the coagulation system, particularly manifested in platelet ultrastructure. It is therefore suggested that platelets may indeed be used successfully to follow the progression of events of cerebral ischemia and possibly assist in the assessment of treatment strategies and their effects on hemostasis. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT We examined CD133 distribution in a human hepatoblastoma cell line (HuH-6 clone 5). We directly observed the cultured cells on a pressure-resistant thin film (silicon nitride thin film) in a buffer solution by using the newly developed atmospheric scanning electron microscope (ASEM), which features an open sample dish with a silicon nitride thin film window at its base, through which the scanning electron microscope beam scans samples in solution, from below. The ASEM enabled observation of the ventral cell surface, which could not be observed using standard SEM. However, observation of the dorsal cell surface was difficult with the ASEM. Therefore, we developed a new method to observe the dorsal side of cells by using Aclar® plastic film. In this method, cells are cultured on Aclar plastic film and the dorsal side of cells is in contact with the thin silicon nitride film of the ASEM dish. A preliminary study using the ASEM showed that CD133 was mainly localized in membrane ruffles in the peripheral regions of the cell. Standard transmission electron microscopy and scanning electron microscopy revealed that CD133 was preferentially concentrated in a complex structure comprising filopodia and the leading edge of lamellipodia. We also observed co-localization of CD133 with F-actin. An antibody against CD133 decreased cell migration. Methyl-β-cyclodextrin treatment decreased cell adhesion as well as lamellipodium and filopodium formation. A decrease in the cholesterol level may perturb CD133 membrane localization. The results suggest that CD133 membrane localization plays a role in tumor cell adhesion and migration. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT To determine whether the geometrical accuracy of small printed elements does not worsen on recycled paper, microscopic analysis of the dot area and the graphic elements raggedness printed on different types of recycled and coated papers at different screen ruling was carried out. Experimental tests have shown that geometrical accuracy of small elements printed on recycled paper/cardboard, in comparison to pictures printed on primary production paper is almost the same. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT In this study, microstructural characterization of Ti-6Al-4V alloy, subjected to the duplex surface mechanical attrition treatment (SMAT)/nitriding treatment, leading to improve its mechanical properties, was carried out through novel and original samples preparation methods. Instead of acid etching which is limited for morphological characterization by scanning electron microscopy (SEM), an original ion polishing method was developed. Moreover, for structural characterization by transmission electron microscopy (TEM), an ion milling method based with the use of two ions guns was also carried out for cross-section preparation. To demonstrate the efficiency of the two developed methods, morphological investigations were done by traditional SEM and field emission gun SEM. This was followed by structural investigations through selected area electron diffraction (SAED) coupled with TEM and X-ray diffraction techniques. The results demonstrated that ionic polishing allowed to reveal a variation of the microstructure according to the surface treatment that could not be observed by acid etching preparation. TEM associated to SAED and X-ray diffraction provided information regarding the nanostructure compositional changes induced by the duplex SMAT/nitriding process. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT The aim of this study was to determine the erosive potential of hydrogen peroxide (HP) containing mouthwash on dentin assessed by Focus variation three-dimensional (3D) microscopy. Twenty dentin slabs were selected and randomly allocated into two groups ( n = 10): DW—Distilled water (pH = 7.27) and HP—1.5% (pH = 3.78). Each specimen was cyclically demineralized (4 × 60 s/day, 10 days) with HP or DW and brushed 3×/day (200 g, 150 strokes—toothpaste with 1,450 ppmF as NaF). Between the challenges, the specimens were exposed to artificial saliva. Afterward, dentin loss was analyzed using focus variation 3D microscopy, and the data were submitted to unpaired t -test (α = 0.05). Statistically significant difference was found between the mean wear rate (μm, ±SD) of HP (1.98 ± 0.51) and DW (1.45 ± 0.39). The results suggest that the use of HP-containing mouthwash associated to brushing may increase the risk of tissue loss and focus variation 3D microscopy may be used as a technique for quantifying dental wear. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT The poultry industry is a sector of agribusiness which represents an important role in the country's agricultural exports. Therefore, the study about embryogenesis of the domestic chicken ( Gallus gallus domesticus ) has a great economic importance. The aim of this study was to evaluate embryonic development of the endoderm in chicken ( Gallus gallus domesticus ). Forty fertilized eggs of domestic chickens, starting from the 1st day of gestation and so on until the 19 days of the incubation were collected from the Granja São José (Amparo, SP, Brazil). Embryos and fetus were fixed in 10% formaldehyde solution, identified, weighed, measured, and subjected to light and scanning electron microscopy. The endoderm originates the internal lining epithelium of the digestive, immune, respiratory systems, and the organs can be visualized from the second day (48 h) when the liver is formed. The formation of the digestive system was complete in the 12th day. Respiratory system organs begin at the fourth day as a disorganized tissue and undifferentiated. Their complete differentiation was observed at the 10 days of incubation, however, until the 19 days the syrinx was not observed. The formation of immune system at 10th day was observed with observation of the spleen, thymus, and cloacal bursa. The study of the organogenesis of the chicken based on germ layers is very complex and underexplored, and the study of chicken embryology is very important due the economic importance and growth of the use of this animal model studies such as genetic studies. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT Lignocellulosic plant cell wall is considered a potential source for second generation biofuels. The plant cell wall is a highly complex structure mainly composed of cellulose, hemicelluloses, and lignin that form a network of crosslinked fibers. The structural organization of the sugarcane cell wall has not been previously analyzed in detail, and this analysis is a prerequisite for further studies on the recalcitrance and deconstruction of its biomass. In this work, cellulose and lignin localization were investigated by confocal laser scanning microscopy. In addition, the internode sugarcane cell wall structural organization was analyzed by electron microscopy. Internode stem anatomy showed a typical monocot structure consisting of epidermis, hypoderm, and vascular bundles scattered throughout ground parenchyma tissue and surrounded by sclerenchyma fibers. Confocal images of safranin labeled sugarcane showed that lignin distribution was predominant in the vessel elements, cell wall corners (CC), and middle lamella (ML), while cellulose-rich cell walls were randomly distributed in the ML and organized in the other cell wall layers. KMnO 4 cytochemistry revealed that lignin was predominantly distributed in secondary cell walls, ML and CC. Cell wall sublayers (S1, S2, and S3) were identified and measured by transmission electron microscopy. Our results provide insights that may help further understanding of sugarcane cell wall organization, which is crucial for the research and technology of plant-based biofuel production. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT This study was undertaken to investigate the pecten oculi of stork by using macroscopic, light and electron microscopic techniques. A total of 20 eyes that were obtained from 10 storks were used. The eyes were cleaned and isolated by dissection. After various procedures, four of the pecten oculi were examined by light microscope while the other four with an electron microscope. The remaining 12 eyes were assigned for macroscopic investigation. Pecten oculi of the stork were determined as accordion-like structures that originated from n. opticus, consisting of 15–17 plica and projecting up to 2/5 of the diameter of the bulbus oculi. Light microscopic examination revealed two types of blood vessels. Afferent–efferent vessels were larger in diamater (40–45 µm), fewer in numbers, and the capillary vessels were smaller in diamater (2–5 µm) and more in numbers. There were granules including amount of melanin pigment at the apical part of the pleats. These granules were fewer and scattered randomly on the basal part of the pleats. As a result, pecten oculi in the stork, which is a migrating bird, were determined to be similar to those of other diurnal birds. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT This study represents the first definitive anatomical description of the tongue and lingual papillae of the pampas deer and compares the different information on the morphology of the other ruminant species available in the literature. In this study, the tongues of four adult and one fetal deer were used. The tongue was elongated with an oval or rounded apex. The filiform papillae on the lingual apex were smaller than the ones on the dorsal and lateral surfaces of the lingual body. Two very thin secondary papillary projections were found to emerge from the bilateral sides of some filiform papillae. Spherical fungiform papillae were randomly distributed among filiform papillae on dorsal surface of the lingual body and ventral surface of the apex. More developed conical papillae were observed in the caudal half of the lingual torus, whereas the rostral half of the torus had smaller conical papillae. Each conical papilla included shallow longitudinal groove on its anterior surface and some conical papillae on the lingual torus had bifid sharp apices. On the caudal portion of the torus, we also observed a few fungiform papillae, which were larger than those located on the lingual body. There were no lenticular papillae on the lingual torus, and five to nine round or oval circumvallate papillae were situated on each caudolateral side of the lingual torus. Morphological features of the tongue in the pampas deer were more similar to wild ruminant species than they were to domestic species. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT Quantitative fluorescence microscopy is severely hindered by intrinsic autofluorescence (AF). Endogenous fluorescent molecules in tissue and cell samples emit fluorescence that often dominates signals from specific dyes. This makes AF removal critical to the development and practice of quantitative fluorescence microscopy. In this study, we showed that AF signal could be separated from specific signal using a customized filter set. The filter set used the same excitation and beam splitter as the standard filter set, but the emission filter was red-shifted 40–60 nm from the peak of the specific dye. This filter set configuration collected mostly AF with minimum contribution from the specific dye. A linear transformation of AF images was required to correct for the difference in exposure and filter configuration. The constants (slope and intercept) in linear transformation were obtained through a pixel to pixel comparison between AF images (no staining) obtained by the standard filter set and the customized AF filter set. After staining of specific dye, the standard filter collecting target dye spectra was used to capture both target signal and AF, whereas customized filter was used to capture only AF. AF removal was accomplished by subtracting the linear transformed AF image from the image obtained from the standard filter. To validate our approach, we examined weak staining of androgen receptor in an AF abundant prostate tissue sample. Our method revealed a similar but cleaner nuclear staining of androgen receptor in a specimen, when compared to a traditional autofluorescence removal method. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT One of the fundamental problems in the analysis of single particle tracking data is the detection of individual particle positions from microscopy images. Distinguishing true particles from noise with a minimum of false positives and false negatives is an important step that will have substantial impact on all further analysis of the data. A common approach is to obtain a plausible set of particles from a larger set of candidate particles by filtering using manually selected threshold values for intensity, size, shape, and other parameters describing a particle. This introduces subjectivity into the analysis and hinders reproducibility. In this paper, we introduce a method for automatic selection of these threshold values based on maximizing temporal correlations in particle count time series. We use Markov Chain Monte Carlo to find the threshold values corresponding to the maximum correlation, and we study several experimental data sets to assess the performance of the method in practice by comparing manually selected threshold values from several independent experts with automatically selected threshold values. We conclude that the method produces useful results, reducing subjectivity and the need for manual intervention, a great benefit being its easy integratability into many already existing particle detection algorithms. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT Tau is a microtubule-associated protein mainly found in neurons. The protein is associated with process of microtubule assembly, which plays an important role in intracellular transport and cell structure of the neuron. Tauopathies are a group of neurodegenerative diseases specifically associated with tau abnormalities. While a well-defined mechanism remains unknown, most facts point to tau as a prominent culprit in neurodegeneration. In most cases of Tauopathies, aggregates of hyperphosphorylated tau have been found. Two proposals are present when discussing tau toxicity, one being the aggregation of tau proteins and the other points toward a conformational change within the protein. Previous work we carried out showed tau hyperphosphorylation promotes tau to behave abnormally resulting in microtubule assembly disruption as well as a breakdown in tau self-assembly. We found that tau's N-terminal region has a putative site for ATP/GTP binding. In this paper we demonstrate that tau is able to bind ATP and not GTP, that this binding induces tau self-assembly into filaments. At 1 mM ATP the filaments are 4–7 nm in width, whereas at 10 mM ATP the filaments appeared to establish lateral interaction, bundling and twisting, forming filaments that resembled the Paired Helical Filaments (PHF) isolated from Alzheimer disease brain. ATP-induced self-assembly is not energy dependent because the nonhydrolysable analogue of the ATP induces the same assembly. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT To overcome the loss of structural integrity when eyeball sections are prepared by wax embedding, we experimentally modified the routine histological procedure and report satisfactorily well-preserved antero-posterior sections of whole eyeballs for teaching/learning purposes. Presently histological sections of whole eyeballs are not readily available because substantial structural distortions attributable to variable consistency of tissue components (and their undesired differential shrinkage) result from routine processing. Notably, at the dehydration stage of processing, the soft, gel-like vitreous humor considerably shrinks relative to the tough fibrous sclera causing collapse of the ocular globe. Additionally, the combined effects of fixation, dehydration, and embedding at 60°C renders the eye lens too hard for microtome slicing at thicknesses suitable for light microscopy. We satisfactorily preserved intact antero-posterior sections of eyeballs via routine paraffin wax processing procedure entailing two main modifications; (i) careful needle aspiration of vitreous humor and replacement with molten wax prior to wax infiltration; (ii) softening of lens in trimmed wax block by placing a drop of concentrated liquid phenol on it for 3 h during microtomy. These variations of the routine histological method produced intact whole eyeball sections with retinal detachment as the only structural distortion. Intact sections of the eyeball obtained compares well with the laborious, expensive, and 8-week long celloidin method. Our method has wider potential usability than costly freeze drying method which requires special skills and equipment (cryotome) and does not produce whole eyeball sections. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT The aim of this study was to evaluate the biocompatibility of mineral trioxide aggregate (MTA) Bio and Portland cement (PC) and compare with those of ProRoot MTA. Polyethylene tubes were filled with materials and placed into dorsal subcutaneous connective tissue of Wistar albino rats. After 7, 30, and 60 days after the surgical procedure, the implants with the surrounding tissue were removed. Tissue samples were subjected to histological processing, and sections were stained with hematoxylin and eosin. Sections were evaluated for the intensity of inflammation, predominant cell type, presence of fibrous capsule and granulation tissue. Data were submitted to Kruskal-Wallis test at a significant level of P ≤ 0.05. No statistical differences were observed at any evaluated condition among tested materials ( P 〉 0.05). Statistically significant differences were observed between mean inflammatory scores, cell types and granulation tissue of the same material in different experimental periods ( P 〈 0.05). Can be concluded that biocompatibility of MTA bio and PC were comparable with that of ProRoot MTA. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: Platelet adhesion is one of the most pivotal events of blood clotting for artificial surfaces. However, the mechanisms of surface-induced platelet activation have not been fully been elucidated or visualized so far. In this study, we attempted to observe the internal structures and adhesion interfaces of human platelets attached to artificial surfaces by transmission electron microscopy (TEM) during the platelet activation process. We prepared observation samples by a conventional embedding method using EPON 812 resin. The sectioning was sliced perpendicular to the a-platelet/material interface. Observation by TEM indicates that internal granules coalesce in the center of the platelet accompanied by pseudopodial growth in the early stage of platelet activation. Pseudopodia from a platelet attach to the material interface not along a plane but at a point. In addition, along with the process of platelet activation, the gap between the platelet membrane and the material surface at the interface disappeared and a-platelet/material adhesion became much tighter. In the fully activated platelet stage, the platelet becomes thinner and tightly adheres to the substrate. As a result of comparative observation of an adherent platelet on polycarbonate (PC) and on amorphous carbon (a-C:H), it was found that internal granules release was inhibited more remarkably on a-C:H coating rather than on PC. Despite numerous technical difficulties in preparing sectional samples, such a study might prove the essential mechanism of biomaterial-related thrombosis, and it might become possible to modify the surfaces of materials to minimize material-related thrombosis. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT In a previous study, the dislocations in Fe 30 Ni 20 Mn 25 Al 25 (at. %), which consist of 50 nm wide alternating b.c.c. and B2 phases, were shown to have a/2〈111〉 Burgers vectors after room temperature deformation. The dislocations were found to glide in pairs on both {110} and {112} slip planes and were relatively widely separated in the b.c.c. phase, where the dislocations were uncoupled, and closely spaced in the B2 phase, where the dislocations were connected by an anti-phase boundary. In this article, we analyze the dislocations in the two ∼5 nm-wide B2 phases in a related two-phase alloy Fe 30 Ni 20 Mn 20 Al 30 , with compositions Fe-23Ni-21Mn-24Al and Fe-39Ni-12Mn-34Al, compressed to ∼3% strain at a strain rate 5 × 10 −4 s −1 at 873 K (the lowest temperature at which substantial plastic flow was observed). It is shown that slip occursby the glide of a 〈100〉 dislocations. A review of the literature suggests that the differences in the observed slip vector between these B2 phases could be due to the differences in composition, differences in deformation temperature, or possibly both. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT The diaphragm is the main respiratory muscle. Along with other respiratory muscles, the diaphragm is responsible for the muscular contraction that generates the respiratory cycle and, as a consequence, the gaseous interchanges in the lungs. Guinea pigs ( Cavia porcellus Linnaeus 1758) are largely used as experimental animals in many biology applications due to their easy management, low cost, and docile behavior. As the diaphragm exerts important effects on lung physiology and function, this study aimed at investigating the morphological characteristics of the muscle, through macroscopic, microscopic, and scanning electron microscopy to add reference data for future studies. We observed a “U”-shaped tendineous center and its morphology was similar to other mammals. These results cooperate with the descriptive and comparative anatomy of mammals, besides can be used as control data for areas of surgery and stem cells. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT The aim of this study was to evaluate the influence of different activation protocols for chelating agents used after chemo-mechanical preparation (CMP), for smear layer (SL) removal. Forty-five single-rooted human premolars with straight canals and fully formed apex were selected. The specimens were randomly divided into three groups depending on the chelating agent used for smear layer removal: distilled water (DW, control group); 17% ethylenediaminetetraacetic acid (EDTA); and 10% citric acid (CA). Each group was further divided into three subgroups according to the activation protocol used: no-activation (NA), manual dynamic activation (MDA), or sonic activation (SA). After CMP, all specimens were sectioned and processed for observation of the apical thirds by using scanning electron microscopy (SEM). Two calibrated evaluators attributed scores to each specimen. The differences between activation protocols were analyzed with Kruskal-Wallis and Mann-Whitney U tests. Friedman and Wilcoxon signed rank tests were used for comparison between each root canal third. When chelating agents were activated, either by MDA or SA, it was obtained the best cleaning results with no significant difference between EDTA and CA ( P 〉 0.05). Sonic activation showed the best results when root canal thirds were analyzed, in comparison to MDA and NA groups ( P 〈 0.05). The activation of chelating agents, independent of the protocol used, benefits smear layer removal from root canals. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT This study investigated the relationship among heat shock protein 70 (HSP70), proliferating cell nuclear antigen (PCNA), and testicular apoptosis during a breeding cycle of Prochilodus argenteus , a neotropical migratory characiform fish of importance in commercial fishery from the São Francisco River basin. A total of 48 (12 fish/sampling) adult males were caught using casting and drifting nets in four samplings from June 2008 to March 2009. Immunohistochemistry, Western blotting, terminal transferase-mediated dUTP nick-end labeling (TUNEL), enzyme-linked immunosorbent assay (ELISA), and caspase-3 colorimetric assay were assessed in different phases of spermatogenesis. Labeling for HSP70 occurred in spermatogonia (SPG A 18.0±1.5 and SPG B 27.9±1.0 in 100 mm 2 , respectively) and Sertoli cells in all sampling periods, with higher values in June (resting period) while spermatocytes were labeled in September (maturation period) and December (ripe period). For PCNA, immunoreaction was predominant in spermatogonia in June and September, while primary spermatocytes were labeled mainly in December (18.7±2.0). TUNEL-positive reaction occurred throughout the sampling periods, and labeling was detected in the nucleus of germ cells in all developmental phases, except spermatozoa. By ELISA, total HSP70 in testis increased significantly from June to December, and decreased in March (regression period), P 〈0.05. Caspase-3 activity decreased from June to December and increased in March. Taken together, our results suggest that HSP70 may protect the germ cells from caspase-3-dependent apoptosis during testicular activity and, reduction of HSP70 and increase of apoptosis contribute for testicular remodeling after the breeding season in wild populations of P. argenteus in the São Francisco River. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT This study aims to describe the morphology of Didelphis sp. male genital organs (penis, testes, epididymis, ductus deferens, prostate, and bulbourethral gland). Ten male animals were used, eight for macroscopic and light microscopy analysis, and two for scanning electron microscopy. The testes and epididymis showed similarity to other eutherian mammals. The bifid penis showed the urethra ending in the medial region where the bifurcation begins, occurring in each segment extension of the urethral groove until the beginning of the glans. Histologically, the penis consists of a cavernous and spongy body, covered by stratified squamous epithelium with loose connective tissue. The urethra was lined by transitional stratified epithelium. In the prostate, prostatic segments were found consisting of tubular glands in a radial arrangement around the urethra, coated externally by a dense connective tissue associated with a relatively thick layer of smooth muscle arranged in two layers that surround the glandular tissue. The animals had three pairs of bulbourethral glands placed at the membranous and cavernous urethra junction with descending and parallel excretory ducts ending caudally in the urethral lumen. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT The objective was to characterize and associate the receptor reactivities of fibroblastic growth factor (FGF)-2, FGF-7, FGF-8, epidermal growth factor (EGF), α-actin and vimentin in relation to the androgen receptor (AR), α and β estrogen receptors (ERα and ERβ), and prolactin receptor in the prostate of elderly men showing low- and high-grade adenocarcinoma. Thirty prostatic samples were taken from 60- to 90-year-old patients without prostatic lesions and with low-grade cancer and high-grade cancer, from the University Hospital, School of Medicine, the State University of Campinas. The results showed that increased FGF-2, FGF-7, and FGF-8 receptor reactivities and decreased AR reactivity were verified in both high- and low-grade cancer. However, the FGF-8 receptor showed greater involvement at the beginning of the malignancy alterations. Increased EGF receptor (EGFR) reactivity and diminished α-actin immunohistochemistry were identified in both cancer groups. Also, increased ERα, PR, and vimentin receptors were verified in both cancer groups. To conclude, the ERα involvement in the reactive stroma activation led to a microenvironment, which was favorable to cancer progression, due to maximizing stromal imbalance. The prolactin could be related to cancer progression due to its interaction with ERα action, indicating that this hormone could be a relevant target to prevent the estrogenic effects in the prostatic lesions. Both FGF receptor (FGFR)-2 and FGFR-8 play a fundamental role in the early stages of prostate cancer, suggesting that these molecules could be a promising therapeutic target. The differential localization of the fibroblastic factors between the prostatic epithelium and stroma of elderly men, who presented prostate cancer, could indicate a favorable distinction for tumoral progression. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT The focused ion beam (FIB) technology has drawn considerable attention in diverse research fields. FIB can be used to mill samples at the nanometer scale by using an ion beam derived from electrically charged liquid gallium (Ga). This powerful technology with accuracy at the nanometer scale is now being applied to life science research. In this study, we show the potential of FIB as a new tool to investigate the internal structures of cells. We sputtered Ga + onto the surface or the cross section of animal cells to emboss the internal structures of the cell. Ga + sputtering can erode the cell surface or the cross section and thus emboss the cytoskeletons quasi-3 dimensionally. We also identified the embossed structures by comparing them with fluorescent images obtained via confocal laser microscopy because the secondary ion micrographs did not directly provide qualitative information directly. Furthermore, we considered artifacts during the FIB cross sectioning of cells and propose a way to prevent undesirable artifacts. We demonstrate the usefulness of FIB to observe the internal structures of cells. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT We describe a simple, effective and relatively cheap instrument for the inflation of soft parts of insect genitalia, particularly the vesica (endophallus) of small male lepidopteran specimens. It can also be applied in female genitalia and for the hyperextension of postabdominal segments of the telescoping type. A fine glass capillary is mounted on a self-constructed micromanipulator system with capillary holder, moveably placed aside a microscope. The micromanipulator is constructed from the x - and y -pinions of a discarded microscope table and the z -pinion of a discarded microscope. Preparation is carried out as usual on a microscopic slide. The inflatable soft parts are pushed out mechanically and then fully inflated and subsequently hardened by pumping absolute alcohol with a medical syringe via a flexible PVC tube, the capillary holder and the glass capillary into the phallus. With affordable effort, our “hands-off”apparatus might considerably ease preparation. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT Many Swertia species are utilized as a traditional medicine under the name “Qingyedan” in China, but are easily confused with one another. To distinguish eight Swertia species ( S. mileensis , S. cincta , S. patens , S. punicea , S. delavayi , S. nervosa , S. macrosperma , and S. yunnanensis ) and to ensure their safety and efficacy, the microscopic and macroscopic characteristics of the roots, stems, leaves, and flowers of them were examined. The results showed that microscopic and macroscopic features helpful for authentication of the eight species were the sinuosity of the anticlinal walls of epidermal cells and presence or absence of hairs on the leaf lamina; presence or absence of V-shaped fibers and fibers with sinuous abaxial wall in the sepals; shape of epidermal cells and pattern of papillae on hairs on the margin of corolla nectary; distribution of stomata in leaf and sepal epidermises, stone cells in cortex and phloem of roots and in cortex and pith of stems, crystals in parenchymatous cells of mesophyll and stem, stomata size, stem diameter, and 4- or 5-merous flowers, and so on. Two keys to the eight Swertia species based on macroscopic and microscopic characteristics are presented. The study indicates that microscopy and related techniques are convenient, practicable, and can be unambiguously applied for authentication of Swertia species. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT Widefield frequency-domain fluorescence lifetime imaging microscopy (FD-FLIM) is a fast and accurate method to measure the fluorescence lifetime, especially in kinetic studies in biomedical researches. However, the small range of modulation frequencies available in commercial instruments makes this technique limited in its applications. Herein, we describe a practical implementation of multifrequency widefield FD-FLIM using a pulsed supercontinuum laser and a direct digital synthesizer. In this instrument we use a pulse to modulate the image intensifier rather than the more conventional sine-wave modulation. This allows parallel multifrequency FLIM measurement using the Fast Fourier Transform and the cross-correlation technique, which permits precise and simultaneous isolation of individual frequencies. In addition, the pulse modulation at the cathode of image intensifier restores the loss of optical resolution caused by the defocusing effect when the cathode is sinusoidally modulated. Furthermore, in our implementation of this technique, data can be graphically analyzed by the phasor method while data are acquired, which allows easy fit-free lifetime analysis of FLIM images. Here, our measurements of standard fluorescent samples and a Föster resonance energy transfer pair demonstrate that the widefield multifrequency FLIM system is a valuable and simple tool in fluorescence imaging studies. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT Among the various corrosion protection strategies for structural steels, coating techniques provide the most cost-effective protection and have been used as the primary mode of corrosion protection. Existing coating techniques however have been used mainly for their barrier capability and therefore all have a limited service life due to oxidation aging, electrolytic degradation, or various inadvertent defects and flaws occurred in and after coating applications. This work investigated the anti-corrosion potential of a π-conjugated polymer—polyaniline (PANi), which was doped into an intrinsically conducting polymer and then included in a two-layer coating system as a primer layer. To achieve a long service life, the primer layer was made by mixing the conductive PANi in a waterborne poly-vinyl butyral solution to provide strong adhesion to steel surface, and then topcoated with a layer of elastomer-modified polyethylene to obtain extra mechanical and barrier protections. Two ASTM standard tests were conducted to evaluate the corrosion durability and tensile adhesion of the two-layer system, in which the system demonstrated superior performance. The Scanning Kelvin Probe Force Microscopy (SKPFM) was used to provide the microscopic evidences for the outstanding performance. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT The sharpness of atomic force microscope (AFM) tips is essential for acquiring high quality AFM images. However, AFM tips would easily get contaminated during scanning and storage at ambient condition, which influences image resolution and causes image distortion. Replacing the probe frequently is a solution, but uneconomical. To solve this problem, several tip cleaning methods have been proposed but there is space for further improvement. Therefore, this article developed a method of tip cleaning by using a one-dimensional grating (600 lines/mm) as a micro-washboard to “wash” contaminated tips. We demonstrate that the contaminants can be scrubbed away by rapidly scanning such micro-washboard against the tip in the aids of Z -dithering (10–20 Hz) exerted on the washboard. This method is highly efficient and proved to be superior to traditional ones. Experiments show that AFM images acquired with “washed” tips have higher resolution and less distortion compared with images acquired using contaminated tips, even comparable to those scanned by new ones. Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.

Description: ABSTRACT The present study evaluated the efficacy of fluazuron (active ingredient of the acaricide Acatak ® ) and its effects on Rhipicephalus sanguineus nymphs fed on rabbits exposed to different doses of this insect growth regulator. Three different doses of fluazuron (20 mg/kg, 40 mg/kg, and 80 mg/kg) were applied on the back of hosts (via “pour on”), while distilled water was applied to the Control group. On the first day of treatment with fluazuron (24 h), hosts were artificially infested with R. sanguineus nymphs. Once fully engorged, nymphs were removed and placed in identified Petri dishes in Biochemical Oxygen Demand (BOD) incubator for 7 days. After this period, engorged nymphs were processed for ultramorphological analysis. The results revealed alterations in the ultramorphology of many chitinous structures (smaller hypostome and chelicerae, less sclerotized scutum, fewer sensilla, fewer pores, absence of grooves, marginal and cervical strips and festoons in the body, even the anal plaque was damaged) that play essential roles for the survivor of ticks and that can compromise the total or partial development of nymphs and emergence of adults after periodic molting. Our findings confirm the efficacy of fluazuron, a more specific and less aggressive chemical to the environment and human health, and that does not induce resistance, in nymphs of the tick R. sanguineus in artificially infested rabbits treated with this arthropod growth regulator (AGR), indicating that it could be used in the control of this stage of the biological cycle of the tick R. sanguineus . Microsc. Res. Tech., 2013 . © 2013 Wiley Periodicals, Inc.