Scanning electron microscopy of melanoblastomas of the choroid

With the scanning electron microscopic study of a case of the fascicular and two cases of epithelioid melanoblastomas it has been found that the shape of the cells is more variable than it was believed before on the basis of light microscopy. Certain types of arrangement described in histological observations, e.g. the fascicular type, can also be well demonstrated by scanning electron microscopy.     

Evaluation of preparation, staining and microscopic techniques for counting incremental lines in cementum of human teeth

Apposition of cementum occurs in phases resulting in two types of layers with different optical and staining properties that can be observed by light microscopy. Narrow, dark staining incremental lines are separated by wider bands of pale staining cementum. The distance from one line to the next represents a yearly increment deposit of cementum in many mammals, and counting these lines has been used routinely to estimate the age of the animals. Incremental lines in cementum have also been observed in sections of human teeth, and the object of the present investigation was to examine a number of methods for preparing and staining them for counting. Longitudinal and transverse sections, either ground or decalcified, were cut from formalin fixed human dental roots, paraffin embedded or frozen, and stained using several techniques. The cementum was investigated using conventional light, fluorescence, polarized light, confocal laser scanning, interference contrast, phase contrast, and scanning electron microscopy. Incremental lines in the cementum could be observed in ground sections and, following decalcification, in both frozen and paraffin embedded sections. Toluidine blue, cresyl violet, hematoxylin, or periodic acid Schiff (PAS) stained incremental lines allowing differentiation by conventional light microscopy. Contrast was best using fluorescence microscopy and excitation by green light since the stained cemental bands, but not the incremental lines, fluoresced after staining with cresyl violet, PAS or hematoxylin and eosin. The results with other microscopic techniques were unsatisfactory. Since incremental lines are not destroyed by acids and stain differently than the remaining cementum, it is likely that they possess an organic structure which differs from the cementum. Incremental lines in human dental cementum could be observed best using decalcified sections stained with cresyl violet excited by green light.     

Incremental lines in root cementum of human teeth: an approach to their ultrastructural nature by microscopy

In ground sections of human teeth, root cementum shows under the light microscope as alternating, almost concentric, dark and light rings. In paleontology and forensic medicine, the number of these incremental lines or annulations is used to derive the age-at-death of the individual. To find the ultrastructural features underlying these cemental annulations, we used bright-field light microscopy (LM), confocal laser scanning microscopy (CLSM), transmission electron microscopy (TEM), and electron-dispersive x-radiation (EDX) in a scanning electron microscope (SEM). Annulations visible in ground sections of about 100-micron thickness were no longer visible in semi-thin sections (thickness, 1-2 microns) of the same specimen in the same area. The assumption that annulations could be caused by super-imposing structures in the depth of field of the light microscope's objective lens was not verified by CLSM. Fiber bundles of higher density than the surrounding matrix in TEM micrographs could not be connected unambiguously with annulations in LM micrographs. After all, the ultrastructural nature of cemental annulations remains an open question.     

A histologic survey of trabeculectomy

Thirty-two tissue blocks removed during trabeculectomy were examined histologically, 16 by both scanning electron microscopy and light microscopy, and 16 by light microscopy alone. Trabecular meshwork was identified in 27 specimens (84.3%) and Schlemm's canal in 17 (53%), each found with equal frequency by either technique. Ciliary muscle was identified in eight of 32 of specimens (25%) examined by light microscopy, but was present in 12 of 16 specimens (75%) examined by scanning electron microscopy. The identification of these specific anatomical structures did not correlate with the postoperative outcome. These findings support the contention that trabeculectomy is a filtering procedure.     

Volume determination of human metaphase chromosomes by scanning force microscopy

The scanning force microscopy (SFM) yields the topography of the investigated surface. A procedure was developed which starts from this three-dimensional information to estimate the volume of a biological specimen. The volume of spread human metaphase chromosomes was determined in air and rehydrated in aqueous buffer. A difference of the determined volume of a air-dried metaphase chromosome set was found compared to values from electron microscopic investigations, and could be correlated with differences in the hydration state of the chromosomes. SFM-based relative volumes of air-dried chromosomes resembles literature data regarding volume range and distribution. Possible application of SFM-based relative volume measurements for chromosome classification purposes is discussed.     

Alterations in nasal sensibility following open rhinoplasty

Scanning electron microscopy (SEM) of methyl methacrylate casts and light microscopy (LM) of tissue are well-established methods for studying the microcirculation. The two are complimentary, but methacrylate is transparent and thus its presence is often not appreciated by LM. Applying histologic stains to sections of tissue embedded in methyl methacrylate would allow the relationships of light microscopic and scanning electron microscopic views of cast vasculature to be better appreciated. We sought to test different stains on cast tissue to find one that would accent the cast. Surgically removed and autopsied human lungs were cast with methacrylate and processed by routine light microscopic methods. They were stained with the hematoxylin and eosin, Masson trichome, elastic--van Gieson, Grocott methenamine silver, Brown-Brennan, and Ziehl-Neelsen methods. The Ziehl-Neelsen procedure stained the methacrylate best, giving it a red color. This procedure also worked well without heating. We conclude that (1) cast methacrylate lung can be processed for routine LM with excellent results; (2) methacrylate stains well with the Ziehl-Neelsen technique; (3) the acid--fast stained cast lung shows capillaries and cells in both normal and diseased lung better than the routine hematoxylin and eosin stain; (4) this technique can be used to assess filling and correlate findings on the same tissue with the two different microscopic methods.     

The evaluation of acrylic resins for the study of nondecalcified human teeth with the light and electronic microscopes

Resin embedding of human teeth for light and transmission electron microscopic studies becomes difficult without previous decalcification. The limited and slow infiltration of the resin into hard tissues may cause problems during preparation and observation of the samples. Moreover the type of resin that is used may affect the morphologic preservation of both tissues and cellular elements. Recently there has been an increasing number of studies on the application of acrylic resins in light and electron microscopy, in order to overcome problems encountered with the use of epoxy resins still utilized in morphologic studies. We compared different acrylic resins (Technovit 7200 VLC, LR White, LR Gold, Bioacryl) in order to understand which one was more suitable for undecalcified human dental tissues under light and transmission electron microscope. Evaluation of such resins was performed using the following criteria: ease of cutting with ultramicrotome, soft and hard tissues infiltration, uptake of tissue stains for both light and electron microscopy, morphologic preservation and stability under electron beam. This study, carried out on the pulp area comprising predentin and dentin, showed excellent quality of Bioacryl and LR Gold, the two resins presenting, by far, the best results among all the different types tested. The optimal morphologic preservation obtained with such resins is indicated for light and electron microscopic studies, allowing their application in different fields of dental research.     

In vitro antioxidant and cytotoxic activity of extracts of Baccharis coridifolia DC

Onchocercal keratitis (river blindness) is one of the leading worldwide causes of blindness. Light microscopic analysis of human specimens and corneal tissue from experimental models has implicated the eosinophil as an important cell in the inflammatory response. Our previous studies in experimental murine onchocercal keratitis have demonstrated that the inflammatory infiltrate is composed primarily of eosinophils displaying ring shaped or bilobed nuclei. However, a number of cells were not characterizable by light microscopy, presumably due to mechanical distortion. To more fully characterize the inflammatory cell infiltrate, we examined corneal specimens by transmission electron microscopy. In addition to typical eosinophils with bilobed and ring shaped nuclei, this approach revealed cells with variable nuclear morphology and cell shape which contained the dense cored granules characteristic of eosinophils. Hence, the degree of pleomorphism of eosinophils is broader than appreciated and underscores the importance of this cell in experimental murine onchocercal keratitis.     

A study on heat source modeling of scanning electron microscopy modified for material processing

Scanning electron microscopy (SEM) has been used as a surface measurement instrument and a tool for lithography in semiconductor processes due to its high-density localized beam. For those purposes, however, the maximum current of SEM is less than 100 pA, which is not enough for material processing. In this article, SEM was modified to increase the amount of current reaching a specimen from the gun part where the current is generated. The maximum current of SEM after modifications was measured at up to 10 μA, which is 10⁵ times greater than before modifications. The current reaching the specimen is called the probe current, which can be defined as a heat source for material processing. The electron beam of the probe current scans rapidly over an area defined by SEM magnification. By considering the electron beam scanning period and the reaction between high-speed electrons and solid material, the heat source model of the rapidly scanning SEM electron beam was suggested. The simulation results with suggested heat source by ABAQUS were compared with the experiment results.     

Scanning and transmission electron microscopy of the spermiophores of Ornithodoros ticks: an attempt to explain their motility

The spermiophores of two tick species, the kangaroo tick, Ornithodoros gurneyi and the cave tick, Ornithodoros tholozani have been examined by scanning and transmission electron microscopy. The anterior end (head) of the spermiophore is a hemisphere covered with a hexagonal network of small projections. The rest of the spermiophore is covered with longitudinal ridges, seen in sections as cellular processes whose membranes are attached only at their anterior ends by specialized 'feet'. In the cytoplasm of the sperm cell body and just beneath the cellular processes are find filaments, which form a continuous layer in O. tholozani and bundles in O. gurneyi. Fibrils tend to be situated beneath the larger cellular processes. In scanning micrographs helical constrictions have been observed in the posterior parts of some spermiophores. It is proposed that certain of the movements observed by light microscopy in living cultures of spermiophores may be explained by contraction of the cytoplasmic filaments seen in the electron microscope.     

A new self-expanding lined stent-graft in the dog ureter: radiological, gross, histopathological and scanning electron microscopic findings

Recurrent or intractable ureteral strictures pose a significant problem for the practicing urologist. Metallic stents have been used sparingly for this problem with varying success. We investigated the use of a stent-graft consisting of a metal stent lined with a porous biocompatible polymer to determine if the liner would prevent urothelial ingrowth. One ureter of each of 11 dogs was treated with either a metallic woven stent or stent-graft inserted retrograde via a midline cystotomy. Six bare wire stents (controls) and five lined with a new, porous, biocompatible, polycarbonate elastomer liner (Corethane) were placed. The animals were followed radiographically with intravenous urography (IVP) at 6 weeks and just prior to sacrifice (12 to 22 weeks). Gross, histological, and electron microscopic analyses were performed. The results demonstrate that all of the bare metal stented animals developed moderate to severe hydroureteronephrosis with significant urothelial hyperplasia and ingrowth through the spaces between the metal wires. The animals implanted with lined stents showed one instance of mild hydroureteronephrosis (observed radiographically but not grossly at time of sacrifice) and virtually no papillary in-growths of urothelium through the stent interstices. This obstructive phenomenon was prevented by the porous polymer lining. There was no evidence of biodegradation of the liner on scanning electron microscopy. Based upon these findings, the marriage of a biocompatible polymer which provokes minimal tissue reaction, and metallic stents which provide tremendous strength, seems to offer significant advantages when placed into the urinary tract to maintain ureteral luminal patency.     

The physical and chemical nature of technegas

Technegas, the discrete radio-aerosol particle, containing 99mTc has been investigated, and the chemical evolution and physical properties of the particle demonstrated. METHODS: A commercial technegas generator was used to produce aerosols according to standard clinical procedures. The aerosols were collected by electrostatic precipitation and examined with transition electron microscopy (TEM), scanning electron microscopy (SEM) and force microscopy. The chemical evolution was examined by x-ray techniques and thermogravimetric analysis. RESULTS: The active particle was identified as hexagonal platelets of metallic technetium contained within a thin layer of graphitic carbon. This composite structure is discussed in light of the metal particle behaving as a template for the carbon capsule. The average size of the observed hexagonal platelets, 30-60 nm, was only weakly dependent on the concentration of technetium in the crucible. CONCLUSION: The mechanism for the formation of the technegas pancreas has been developed and the particles involved characterized. It appears that the use of other metals also leads to the formation of similar materials.     

Pulmonary alveolar septal amyloidosis. A scanning- and transmission electron microscopy study

A case of primary amyloidosis is presented with diffusely involved the alveolar septa. The lung was studied by light microscopy and by transmission and scanning electron microscopy. The fine structure of the amyloid material showed it to be porous, homogeneous, and an acellular substance consisting of interwoven bundles of amyloid fibrils. The fine structure of the amyloid material was considered to explain the normal gas diffusion across the alveolar respiratory membrane. The diagnosis of amyloidosis was first made from a uterine cervical biopsy specimen.     

Intraepidermal mass of M. leprae in a case of seborrheic keratosis due to Hansen's disease (LLs)

A 67-year-old patient has had exanthema in the lower right limb since 51 years ago (16 years old at onset), which underwent repeated remission and recurrence. At present, he has bilateral symmetrical widespread infiltrating exanthema and asymmetrical marked neuralhypertrophy, and has been diagnosed typical LLs (His father had the same disease). The exanthema recurred several years ago, and the patient is being treated for Hansen's disease. He had a dark brown flat elevation with a rough surface and the size of a small finger tip in his right abdominal skin for approximately 20 years. A biopsy was performed, and the specimen was fixed in 10% formalin and paraffin sections were prepared for histopathologic examination. A part of the specimen was processed forscanning electron microscopic examination. Seborrheic keratosis was diagnosed by H & E staining. Acid-fast (FITE) staining, immunohistochemical staining (keratin, S-100 protein, anti-PGL antibody and anti-BCG antibody) and scanning electron microscopy revealed the presence of bacteria (M. leprae) in the dermal foam cells, the matrix with a banded structure and the squamous epithelial cells which normally lack phagocytosis function. Compared to the basal cells of normal epidermis, the basal cells located adjacent to the dermis affected with seborrheic keratosis showed increased proliferation and more marked characteristics of a germinative cell. The degree of differentiation of the basal cells appeared regressed, and they probably possessed augmented phagocytic activity. The phagocytosed bacteria were probably carried by the epidermal cell cycle toward the surface layer. However, bacteria could not be found in the stratum corneum, probably due to an association with the lysosome.     

Role of electron microscopy in transplant renal pathology

The crucial role that electron microscopy plays in diagnostic renal pathology is undisputed. By allowing recognition of findings not identifiable by light microscopic evaluation, electron microscopy has contributed significantly to the understanding of renal diseases and has proven to be of unquestionable value in many diagnostic situations. However, the percentage of cases in which electron microscopic examination adds important information that is either key for establishing or confirming a diagnosis or provides valuable data that influence patient's management remains controversial. This figure depends on the renal biopsy service that is surveyed, but it is reported that on the average ultrastructural evaluation is of value in approximately 30 to 45% of the cases. Correct interpretation of a renal biopsy depends on the ability to correlate light, immunofluorescence, and ultrastructural findings. In contrast, the role of electron microscopy in the examination of renal transplant specimens remains controversial. Many centers do not use routine electron microscopy to examine these specimens and insist that there are only a few specific indications that require ultrastructural evaluation. There is general agreement among renal pathologists that electron microscopy is of importance in the evaluation of renal specimens from patients with proteinuria to distinguish between transplant glomerulopathy, recurrent or de novo glomerulonephritis in order to correctly manage these patients and predict survival of the graft. The other possible indications are much more controversial. This paper summarizes and critically reviews the literature available on this subject and defines recommendations based on the information available at the current time.     

Evaluation of nerve recovery from minimal-duration crush injury

The recovery of damaged peripheral nerves has been the subject of multiple studies. The effects of an inadvertent clamping of a nerve has not been well examined. An experiment was performed to evaluate the effects of a minimal-duration crush injury on the rat sciatic nerve and to determine if walking track analysis was useful in evaluating the short-term functional deficit. Ten Sprague-Dawley rats underwent high-pressure, short-duration crush injuries. Walking track analysis was done regularly for 3 weeks. Histological specimens for light and electron microscopy were taken at postoperative days 3, 7, 14, 21, and 42 from similar animals. There was significant decrease in function by the second week, which then improved to control levels after week 3. Toluidine blue and electron microscopic findings confirmed the clinical course, while routine histological findings tended to lag behind the return of function. Walking track analysis appears to be an effective method of evaluating the short-duration nerve crush injury.     

Microstructural analysis of vanadium carbide/steel surface-alloyed materials fabricated by high-energy electron-beam irradiation

This study is concerned with the microstructural analysis of vanadium carbide (VC)/steel surface-alloyed materials fabricated by high-energy electron-beam irradiation. The mixtures of VC powders and MgO-CaO flux were deposited on a plain carbon steel substrate, and then electron beam was irradiated on these mixtures using an electron-beam accelerator. Microstructures of the irradiated surface regions were examined by optical microscopy, scanning electron microscopy, and transmission electron microscopy. Residual pores were found in the specimen processed without flux, but hardly found in the specimens processed with a considerable addition of flux. As a result of irradiation, vanadium content was homogeneously maintained throughout the melted region, and fine vanadium carbides were formed in the melted region. These microstructural modification including the formation of vanadium carbides greatly improved hardness, especially high-temperature hardness up to 500 °C.     

Ultrastructural and immunohistochemical studies of intralobular fibroblasts in human submandibular gland: the recognition of a 'CD34 positive reticular network' connected by gap junctions

In order to clarify the ultrastructural and immunophenotypic characteristics of intralobular stromal cells in the human submandibular gland, normal tissues were examined from 25 patients by conventional histology of wax sections, light microscope immunohistochemistry, conventional thin-sectioning transmission electron microscopy, immuno-electronmicroscopy (for the CD34 antigen) and scanning electron microscopy. Spindled stromal cells had abundant rough endoplasmic reticulum cisternae typical of fibroblasts. Some actin filaments and microtubules were seen, but the filaments lacked the focal densities of smooth-muscle, nor was a lamina present. Fibroblasts exhibited long slender processes which associated with other fibroblasts by means of gap junctions and with mononuclear cells (lymphocytes, plasma cells, macrophages) by means of close appositions of membrane. Immunoelectron microscopy for CD34 revealed positive staining over the fibroblast surfaces but not over the mononuclear cells. The novel observation of gap junctions suggests a tissue closely organized as a network in physical and perhaps physiological terms. This network also incorporates mononuclear cells. The possible role in immunosurveillance of this CD34-positive reticular network is discussed.     

A method of measuring stored energy macroscopically using statistically stored dislocations in commercial purity aluminum

Stored energy from plastic deformation in rolled aluminum has been quantified with both macroscopic and microscopic methods. Differential scanning calorimetry (DSC) and Microhardness tests were used to determine a value for stored energy based on energy released during recrystallization and resistance to plastic flow from the accumulated dislocation content, respectively. For a value of stored energy based only on geometrically necessary dislocations, orientation imaging microscopy (OIM) within a scanning electron microscope (SEM) was used and supported by transmission electron microscopy (TEM) observation of subgrain cell structure. A value for the average misorientation angle that could be associated with the TEM was obtained from the OIM data. The values of stored energy derived from the various analyses were found to be similar with slight overestimation from the OIM technique. Thus, the difference between the macroscopic and microscopic methods represented the statistically stored dislocations. This article is based on a presentation made in the symposium entitled “Processing and Properties of Structural Materials,” which occurred during the Fall TMS meeting in Chicago, Illinois, November 9–12, 2003, under the auspices of the Structural Materials Committee.     

Microstructures induced by a stress gradient in a nickel-based superalloy

The evolution of the microstructure of single crystals of a nickel-based superalloy during high temperature (1323 K, 1050°C) creep in bending has been studied. Bending provides both tensile and compressive stress gradients; consequently the effects of varying stress conditions on the evolution of the morphology of the γ′ precipitates can be determined from a single specimen. The morphological changes were analysed by scanning electron microscopy using image analysis techniques and by transmission electron microscopy, then described by dimensionless parameters. We discuss the dependence of the morphological changes in the superalloy on the stresses acting in the sample (magnitude and sign). We also discuss the driving mechanisms for the observed morphological changes.