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PubMed2004

Analysis of environmental particles by atomic force microscopy, scanning and transmission electron microscopy.

Due to their large specific surface and their abundance, micro and nano particles play an important role in the transport of micropollutants in the environment. Natural particles are usually composed of a mixture of inorganic amorphous or crystalline material (mainly FeOOH, Fe(x)Oy, Mn(x)Oy and clays) and organic material (humics and polysaccharides). They all tend to occur as very small particles (1-1,000 nm in diameter). Most natural amorphous particles are unstable and tend to transform with time towards more crystalline forms, either by aging or possibly, by dissolution and re-crystallization. Such transformations affect the fate of sorbed micropollutants and the scavenging properties are therefore changed. As these entities are sensitive to dehydration (aggregation, changes in the morphology), it is highly important to observe their morphology in their natural environment and understand their composition at the scale of the individual particles. Also for the understanding and optimization of water treatment technologies, the knowledge of the occurrence and behavior of nano-particles is of high importance. Some of the possible particle analysis methods are presented: aggregation processes, biomineralization, bacterial adhesion, biofilms in freshwaters, ferrihydrite as heavy metals remover from storm water. These examples demonstrate the capabilities and focus of the microscopes. Atomic Force Microscopy (AFM) allows to analyze the particles in their own environment, meaning in air or in the water. Thus, native aspects of particles can be observed. As well, forces of interactions between particles or between particles and other surfaces such as membranes will be highly valuable data. Scanning Electron Microscopy (SEM) and for higher lateral resolution, Transmission Electron Microscopy (TEM) allow measurement of the morphology and composition. Especially, TEM coupled with Electron Energy Loss Spectroscopy (TEM-EELS) is a powerful technique for elemental analysis. Finally, general guidelines for the effective use of microscopic techniques are provided.

Bacteria↗
PubMed1986

Characterization of mucus glycoconjugates in normal human conjunctiva by lectins in light microscopy, transmission and scanning electron microscopy.

Maintenance of tear film in normal conditions is dependent on mucus layer integrity and the presence and distribution of conjunctival epithelial cell microvilli. In the present work a new methodology has been developed to gain correlative information about microprojection assessment and mucus composition, from the same specimen, by Light Microscopy (LM), Transmission Electron Microscopy (TEM) and Scanning Electron Microscopy (SEM). We have characterized the glycosidic residues secreted by goblet cells in normal human conjunctiva, by means of four lectins (WGA, ConA, PNA and SBA), conjugated with FITC for LM and with colloidal gold for TEM and SEM. The cytochemical reactions were performed on histological sections of paraffin-embedded material and on semithin and ultrathin sections of both Epon embedded material directly processed for TEM and of blocks recovered from SEM and reprocessed for TEM. WGA, ConA, PNA and SBA receptors were found to be constituents of the mucus produced by goblet cells in human conjunctiva. The granules of the so-called "second mucus system" (SMS) cells were labelled mainly by WGA. A difference in the quality of glycoconjugates between goblet cells and SMS cells has been also demonstrated. Our results provide an improved method to evaluate alterations of tear film that occur in many conjunctival diseases.

Carbohydrates↗
PubMed1983

Movement of cell surface immunoglobulin (Ig) in guinea pig B cells and lymphocytic leukemia cells as observed by light microscopy and scanning and transmission electron microscopy.

The distribution and regeneration of immunoglobulin (Ig) of guinea pig leukemia cells were investigated through the use of ferritin labeling and scanning electron microscopy. Throughout this work, correlative light microscopy using fluorescein label and transmission electron microscopy using ferritin label were used. The cells used in this study were lymphocytic leukemia cells, an acute (L2C) and a chronic (KSL) form, and normal B cells obtained from Sewall Wright strain 2 guinea pigs. Distinct differences in the movement and regeneration of cell surface Ig were observed when these cells were compared. Both L2C and KSL cells were slower to cap than normal B cells which formed a well-organized single patch. The cells endocytosed the label rapidly and either processed or shed the label within 24 hours except for the L2C cells which had retained internalized label after 24 hours. Regeneration of surface Ig was clearly demonstrated in all three cell types. The apparent similarities between these two lymphocytic guinea pig leukemias and similar reports of human leukemias strongly suggest that the L2C and KSL cells could provide excellent models for future studies of acute and chronic forms of this disease.

Animals↗
PubMed1983

Scope of scanning electron microscopy, transmission electron microscopy and freeze fracture technique in diagnostic cytology of effusions.

A review of the literature on the application of scanning electron microscopy (SEM) and transmission electron microscopy (TEM) to cytodiagnosis of neoplastic cells in effusion fluid reveals a lack of consensus as to which of the techniques is most beneficial to a cytologist for a satisfactory diagnosis. The purpose of this study is therefore to critically evaluate the relative merits of SEM, TEM and freeze-fracture (FF) techniques in enhancing the diagnostic "sensitivity" of ultrastructural cytology. Two cases of adenocarcinoma, one case of mesothelioma and one case of small cell undifferentiated carcinoma of the lung were studied in detail by SEM, TEM and FF techniques. A comparative analysis of the results demonstrates that SEM is a rapid and useful technique where the microvilli on the tumour cell surface is the most prominent feature especially in adenocarcinomas. The major handicap of SEM becomes evident in cases where the extreme variability of the surface morphology amongst tumour cells makes it difficult to establish a diagnosis with confidence. TEM along with light microscopy (LM) of toluidine blue sections gives us the most comprehensive method for cyto-diagnosis and is particularly useful in cases where LM cytology alone is inconclusive. It has the added advantage of revealing diagnostically significant nuclear and cytoplasmic features which are helpful in differentiating a benign from a malignant cell and in some instances the probable site of the primary neoplasm. FF technique, though not conducive to a routine diagnostic situation, is of benefit in demonstrating the organization of junctional complexes. From preliminary studies it appears that the tight junctions in adenocarcinoma are different from those seen in small cell carcinoma. In mesothelioma, gap junctions are invariably present, but they are not seen prominently in adenocarcinoma or small cell carcinoma. Thus in poorly differentiated tumours where LM and TEM are inconclusive, FF may prove useful. It is concluded that such coordinated studies in the future have the potential to better our understanding of the nature and behaviour of tumour cells in effusion fluid.

Adenocarcinoma↗
PubMed1993

[Detecting respiratory syncytial virus in respiratory epithelial cells in adult patients by electron microscopy and immune electron microscopy].

In order to evaluate the value of electron microscopy (EM) in diagnosing respiratory syncytial virus (RSV) infection in adults, the exfoliated cells from nasopharynx of 97 RSV positive patients were examined by EM and the gold labelling technique for immune electron microscopy. The viral particles with RSV features were found at the surface of the exfoliated cells only in a few samples by EM, and the specific gold labelling were observed in the immune electron microscopy (IEM). The results showed that RSV is one of the causative agents for adult respiratory infection.

Adolescent↗
PubMedDate not supplied

[A comparison between phase-contrast optical microscopy and scanning electron microscopy for the analysis of air-borne asbestos fibers in an office environment].

There is no general agreement on the relationship between results obtained by optical microscopy (PCOM) and electron scanning microscopy (SEM) or electron transmission microscopy (TEM) for airborne asbestos fibres. However, a considerable amount of data has been produced in latter years indicating that a general correlation factor, not differing greatly from unity, can be established between the two analytical techniques if counts are limited to fibres longer than 5 microns. A study was made of the relationship between SEM and PCOM when used simultaneously for determination of background concentrations of asbestos in office buildings. Twenty-four couples of samples were collected in parallel for PCOM and SEM in different locations distributed over the various floors of two buildings containing amosite sprayed on structural parts. Asbestos and total fibre counts by SEM were performed following the Verein Deutscher Ingenieure (VDI) method 3492. For total fibre counts the European PCOM method was mainly used. Discrimination of asbestos from non-asbestos fibres was performed using criteria based on birefringence and morphological characteristics of fibres. Counts included only those fibres longer than 5 microns, diameter less than 3 microns and aspect ratio greater than 3. Both total fibres and asbestos fibres were recorded separately by PCOM and SEM. the arithmetic and geometric means of the ratio between PCOM and SEM were 1.3 and 1.1 respectively for total fibres and 1.2 and 1.1 for asbestos fibres (excluding one outlying value). The regression study showed a significant linear correlation (P = 0.01) with correlation coefficients of 0.84 for asbestos fibres and 0.52 for total fibres.

Air Pollution, Indoor↗
PubMed1995

Examination of injected specimens by confocal laser scanning microscopy and scanning electron microscopy.

Although corrosion casts commonly examined by scanning electron microscopy (SEM) offer good insights into the general arrangement of the microvasculature of certain organs, no information can be obtained from such specimens on special morphological data or dynamic events. Thus, most investigators are obliged to make their interpretations of cast structures only on the basis of indirect criteria. A synopsis, which considers the most important data in that respect, is given for the blood and lymphatic microvessels. Moreover, a methodological approach is introduced, based on the application of fluorescent resin, which allows the examination of cast structures in uncorroded specimens together with the surrounding tissue by confocal laser scanning microscopy (CLSM). When two differently stained resins are injected at different sites, e.g., Mercox-fluorescent yellow into an artery and Mercox-rhodamine into the interstice, blood vessels and lymphatics can be well distinguished. In lymph nodes, the system of lymphoid spaces and that of intranodal sinuses can be separately represented with the same technique. Applications and advantages of the CLSM method, which is a useful tool for an extended interpretation of corrosion casts in SEM, are shown in cast specimens from tongue, skin, and liver in rats. In this study, both corroded and uncorroded specimens were considered. In addition, a new approach is demonstrated which combines casting technique and fluorescence microscopy with investigations carried out under experimental conditions. This allows the examination of microvascular casts together with labelled cells in liver and spleen in a state of phagocytosis.

Animals↗
PubMed1979

Hormonal influences on the dog prostate--a correlation study with light microscopy, transmission and scanning electron microscopy.

Dog prostates under defined hormonal influences were investigated by light microscopy (LM), transmission electron microscopy (TEM) and scanning electron microscopy (SEM). The normal acinar cell apices exhibited well-developed, densely packed microvilli. TEM demonstrated numerous secretory granules, indicating a high secretory activity. Following castration, the acinar cells of the dog prostate showed rounded apices. Most of the microvilli disappeared and only knob-like protrusions were seen. Castrated dogs following administration of 3 alpha-androstandiol showed fewer microvilli than the controls; however, a well-developed undulating surface could be observed. Castrated dogs given 3 alpha-androstandiol and cyproterone acetate showed almost completely atrophic glandular cells. Prostates of dogs with spontaneous benign prostatic hyperplasia (BPH) were characterized by pleomorphism of cell shapes and surfaces. The different hormonal influences induced changes of the prostatic surface structures and secretory activity.

Androstanols↗
PubMed1998

Cytoskeletal regulation of human salivary secretion studied by high resolution electron microscopy and confocal laser microscopy.

To study the cell regulation mechanisms of human salivary secretion, surgical specimens of human parotid and submandibular glands were treated in vitro with isoproterenol (beta-agonist), carbachol (muscarinic agonist), and cytochalasin D (microfilament disruptive agent), and morphological changes occurring in serous acinar cells were observed. Control acinar cells treated without secretagogues exhibited only occasional examples of exocytosis. Microfilaments, revealed by transmission electron microscopy (TEM) and confocal laser scanning microscopy (CLSM) of F-actin fluorescence stained by rhodamine-phalloidin, were localized underneath the luminal membrane to separate the secretory granules from the luminal membrane. Following isoproterenol treatment, secretory granules made direct contact with the luminal membrane and many omega-shaped exocytotic profiles appeared. TEM and scanning electron microscopy (SEM) showed these profiles to be of granule size or somewhat smaller and to be provided on their cytoplasmic surface with coated pits. Furthermore, CLSM detected the appearance of F-actin fluorescence around the exocytosed granule membranes. Carbachol treatment also evoked the formation in acinar cells of omega-shaped exocytotic profiles some of which were larger than the granules and which exhibited neither coated pits nor associated F-actin fluorescence. To determine if microfilaments regulate the post-exocytotic process of membrane retrieval, we combined isoproterenol treatments with cytochalasin D or carbachol. Following these treatments, F-actin fluorescence surrounding the exocytosed membrane was dispersed or diffused and the exocytotic profiles enlarged remarkably. These results led to the hypothesis that exo/endocytotic processes in human salivary serous acinar cells are regulated differently under autonomic receptor control mediated by microfilaments.

Adult↗
PubMed2004

Imaging microscopy of the middle and inner ear: Part I: CT microscopy.

Anatomic definition of the middle ear and bony labyrinth in the clinical setting remains limited despite significant technological advances in computed tomography (CT). Recent developments in ultra-high resolution imaging for use in the research laboratory on small animals and pathologic specimens have given rise to the field of imaging microscopy. We have taken advantage of this technique to image a human temporal bone cadaver specimen to delineate middle ear and labyrinthine structures, only seen previously using standard light microscopy. This approach to the study of the inner ear avoids tissue destruction inherent in histological preparations. We present high-resolution MicroCT images of the middle ear and bony labyrinth to highlight the utility of this technique in teaching radiologists and otolaryngologists clinically relevant temporal bone anatomy. This study is not meant to function as a complete anatomic atlas of the temporal bone. We have selected several structures that are routinely delineated on clinical scanners to highlight the utility of imaging microscopy in displaying critical anatomic relationships in three orthogonal planes. These anatomic relationships can be further enhanced using 3D volume rendering.

Cadaver↗
PubMed1997

The fine structure of fenestrated adrenocortical capillaries revealed by in-lens field-emission scanning electron microscopy and scanning transmission electron microscopy.

Cell biologists probing the physiologic movement of macromolecules and solutes across the fenestrated microvascular endothelial cell have used electron microscopy to locate the postulated pore within the fenestrae. Prior to the advent of in-lens field-emission high-resolution scanning electron microscopy (HRSEM) and ultrathin metal coating technology, quick-freeze, platinum-carbon replica and grazing thin-section transmission electron microscopy (TEM) methods provided two-dimensional or indirect imaging methods. Wedge-shaped octagonal channels composed of fibrils interwoven in a central mesh were depicted as the filtering structures of fenestral diaphragms in images of platinum replicas enhanced by photographic augmentation. However, image accuracy was limited to replication of the cell surface. Subsequent to this, HRSEM technology was developed and provided a high-fidelity, three-dimensional topographic image of the fenestral surface directly from a fixed and dried bulk adrenal specimen coated with a 1 nm chromium film. First described from TEM replicas, the "flower-like" structure comprising the fenestral pores was readily visualized by HRSEM. High-resolution images contained particulate ectodomains on the lumenal surface of the endothelial cell membrane. Particles arranged in a rough octagonal shape formed the fenestral rim. Digital acquisition of analog photographic recordings revealed a filamentous meshwork in the diaphragm, thus confirming and extending observations from replica and grazing section TEM preparations. Endothelial cell pockets, first described in murine renal peritubular capillaries, were observed in rhesus and rabbit adrenocortical capillaries. This report features recent observations of fenestral diaphragms and endothelial pockets fitted with multiple diaphragms utilizing a Schottky field-emission electron microscope. In-lens staging of bulk and thin section specimens allowed tandem imaging in HRSEM and scanning TEM modes at 25 kV.

Adrenal Cortex↗
PubMed1975

Scanning electron microscopy of the wall of the third ventricle of the brain of Rana temporia. II. Electron microscopy of the ventricular surface of the pars ventralis of the tuber cinereum.

Scanning electrom microscopy of the ventricular surface of the pars ventralis of the tuber cinereum of the frog Rana temporaria shows that it can be divided in (1) a dorsolateral area, characterized by the presence of very numerous large, solid, bulbous protrusions (2) a medial area, where the large bulbous structures are completely absent and which is diffusely covered with very numerous cilia, (3) a transiotional area located between the two other regions. The large bulbs, shown by scanning electron microscopy, correspond with the end-bulbs of thick ventricular dendrites of nerve cells revealed by transmission electron microscopy. At least many of these intra-ventricular bulbs are dendritic endings of peptidergic neurosecretory neurons, which have been tentatively identified as adenohypophysiotropic neuro-hormone producing neurons (Derickx et al., 1972, 1973a, 1973b). The structural features of the dendritic endings plead in favour of a possible receptive role.

Animals↗
PubMed1984

Granular cell tumour of the mammary gland simulating malignancy. A report on two cases with light microscopy, transmission electron microscopy and immunohistochemical investigation.

Primary granular cell tumours of the breast in 35 and 55 year old women were studied by light microscopy, transmission electron microscopy and immunohistochemistry. Light and electron microscopy revealed a neural origin of the tumours and this was further substantiated by immunohistochemical studies, with positive S-100 protein reaction and negative reactions for surface heavy and light chains, CEA, alfa-1-antitrypsin, muramidase and GFA-protein. Granular cell tumour of the mammary gland is a very rare tumour. Clinically it sometimes simulates carcinoma because of its fibrous consistency, fixation to pectoral fascia and skin retraction. The diagnosis of granular cell tumour should be included in the differential diagnosis of carcinoma of the breast. The granular cell tumour is derived from neuro-ectodermal tissue. Whether it represents a neurogenic cell-confined metabolic disturbance with lysosomal activation, or a true neoplasm remains to be elucidated.

Adult↗
PubMed1982

Comparison of solid-phase immune electron microscopy, direct electron microscopy and enzyme-linked immunosorbent assay for detection of rotavirus in faecal samples.

One hundred and fifteen faecal samples from children with gastroenteritis have been examined by solid-phase immune electron microscopy (SPIEM), standard direct electron microscopy and enzyme-linked immunosorbent assay (ELISA). Diluted high-titered anti-(human) rotavirus serum, produced in rabbit by immunization with purified pooled rotaviruses from gastroenteritis patients, was used as antiserum in the immunological tests. The presence of rotavirus was demonstrated in 41 (36%) of the 115 samples by both SPIEM and ELISA. No false positive reactions were seen. By direct electron microscopy rotavirus was detected in 35 (30%) of the specimens. In addition, other gastroenteritis viruses like adeno-, astro- and calicivirus were visualized by this non-specific test in eight samples. The applicability of these three methods in routine diagnostic work is discussed.

Antigen-Antibody Reactions↗
PubMed1993

Amplified surface microscopy. Preliminary evaluation of a 400-fold magnification in the surface microscopy of cutaneous melanocytic lesions.

BACKGROUND: Surface microscopy (SM) opened a new dimension in the clinical assessment of cutaneous pigmented lesions. Diagnostic patterns were described to provide guidelines for the preoperative diagnosis of pigmented skin lesions. OBJECTIVE: Our purpose was to explore whether "amplified" surface microscopy (ASM), by increasing magnification up to 400-fold, provides any improvement in the analysis of patterns previously described in cutaneous pigmented lesions. METHODS: A fiber-optic camera, allowing magnification up to 400 times and associated with an imaging system computer, was used for the analysis of 40 melanocytic lesions. Special emphasis was put on the most important features that can be observed in surface microscopy, the so-called pigment network and brown globules. RESULTS: The lines (grids) of the pigment network appeared to be composed of three zones: two darker external lines and one central clearer zone. This pattern, not previously seen with SM, is compatible with the anatomy of pigment accumulation in the epidermal rete ridges. Two types of brown globules could be distinguished: one located in the center of the pigment network holes, the other on the lines of its grids. The "on-the-grid" type was smaller than the "in-the-hole" type; it could be misinterpreted as "black dots" when seen at lower magnification (x10). CONCLUSION: ASM is a tedious procedure that may help in the analysis of the features previously recognized by SM; SM is more accessible for routine use.

Diagnosis, Differential↗
PubMed1985

Scanning electron microscopy and transmission electron microscopy of the ciliated cells of the trachea of the rabbit treated with misonidazole alone and in combination with ionizing radiation.

The trachea is often located in the treatment volume when irradiating malignant tumours in the thorax. In order to evaluate possible synergism between misonidazole and irradiation on this tissue, the following studies were made. Fifty rabbits were treated with daily injections of 100 mg misonidazole given i.p. on consecutive days from 1 to 10 days. Morphological investigations of the trachea were made with scanning electron microscopy (SEM), transmission electron microscopy (TEM) and light microscopy (LM). Physiological examinations were performed with recording of the ciliary beat frequency. The results were compared with those from a group of 100 rabbits given misonidazole in a similar manner and exposed to irradiation (2 Gy) 15-30 min after each injection. Ten rabbits were used as controls. The results are compared to the effect of fractionated irradiation alone with 2 Gy/day. Fractionated irradiation of the ciliary epithelium in the trachea of the rabbit has shown dose-dependent physiological and morphological effects. Misonidazole potentiates these effects of radiation with a more pronounced change of the ciliary beat frequency and an increased metabolic activity as could be visualized on TEM. The combination of drug and irradiation also induced a hyperplasia of the ciliary epithelium. Misonidazole itself had no effect on the ciliary beat frequency, but caused a hypoplasia of the ciliary epithelium.

Animals↗
PubMed2006

Characterization of acrylic polyamide plastic embolization particles in vitro and in human tissue sections by light microscopy, infrared microspectroscopy and scanning electron microscopy with energy dispersive X-ray analysis.

Vascular embolization is a well-established practice for the treatment of tumors and vascular lesions. Rounded beads (microspheres) of various materials (collagen, dextran and trisacryl-polymer-gelatin) were developed to solve problems encountered with earlier versions of embolic material. We performed histochemistry, Fourier transform infrared microspectroscopy and scanning electron microscopy with energy dispersive X-ray analysis on two uterine and one hepatic specimen with unidentified intravascular foreign material, and examined a reference embolization product for comparison. The hematoxylin and eosin stained tissue sections showed multiple foci with unidentified intravascular foreign material and fibrous obliteration of vessel lumens. Only one case had a clinical history of previous embolization but without specifying the material used. One case was submitted for identification of a 'parasite'. The material stained positively with Sirius red and mucicarmine, variably with Masson's trichrome stain and Movat pentachrome, and did not stain centrally with periodic acid Schiff with diastase. Infrared spectrophotometric analysis of the material from all three cases demonstrated the spectrum of acrylic polyamide plastic. A control sample of EmboGold exhibited infrared microspectroscopic spectra similar to the three tissue specimens. Analysis by scanning electron microscopy with energy dispersive X-ray analysis demonstrated some differences in elemental composition between the tissue sections and the selected reference material. To our knowledge, this is the first report of infrared spectrophotometric analysis with scanning electron microscopy with energy dispersive X-ray analysis of an acrylic polyamide plastic embolization product both in vitro and in human histologic tissue sections. In cases lacking appropriate clinical information, identification by these methods and/or a panel of special stains may assist pathologists unfamiliar with this material's light microscopic appearance.

Acrylic Resins↗
PubMed1997

Comparison of two-photon excitation laser scanning microscopy with UV-confocal laser scanning microscopy in three-dimensional calcium imaging using the fluorescence indicator Indo-1.

Two-photon excitation laser scanning fluorescence microscopy (2p-LSM) was compared with UV-excitation confocal laser scanning fluorescence microscopy (UV-CLSM) in terms of three-dimensional (3-D) calcium imaging of living cells in culture. Indo-1 was used as a calcium indicator. Since the excitation volume is more limited and excitation wavelengths are longer in 2p-LSM than in UV-CLSM, 2p-LSM exhibited several advantages over UV-CLSM: (1) a lower level of background signal by a factor of 6-17, which enhances the contrast by a factor of 6-21: (2) a lower rate of photobleaching by a factor of 2-4; (3) slightly lower phototoxicity. When 3-D images were repeatedly acquired, the calcium concentration determined by UV-CLSM depended strongly on the number of data acquisitions and the nuclear regions falsely exhibited low calcium concentrations. probably due to an interplay of different levels of photobleaching of Indo-1 and autofluorescence, while the calcium concentration evaluated by 2p-LSM was stable and homogeneous throughout the cytoplasm. The spatial resolution of 2p-LSM was worse by 10% in the focal plane and by 30% along the optical axis due to the longer excitation wavelength. This disadvantage can be overcome by the addition of a confocal pinhole (two-photon excitation confocal laser scanning fluorescence microscopy), which made the resolution similar to that in UV-CLSM. These results indicate that 2p-LSM is preferable for repeated 3-D reconstruction of calcium concentration in living cells. In UV-CLSM, 0.18-mW laser power with a 2.6-phi pinhole (in normalized optical coordinate) gives better signal-to-noise ratio, contrast and resolution than 0.09-mW laser power with a 4.9-phi pinhole. However, since the damage to cells and the rate of photobleaching is substantially greater under the former condition, it is not suitable for repeated acquisition of 3-D images.

Animals↗