Fiber analysis of an insect peripheral nerve: Light vs. electron microscopy

This paper describes the morphology of one of the two major femoral nerve trunks (nerve V of Dresden and Nijenhuis (1958)) of the cockroach Blaberus discoidalis as revealed by light microscopy, scanning electron microscopy and transmission electron microscopy. Fiber analysis of light and electron micrographs of thick and thin sections taken from adjacent areas of the same nerve yield significantly different data about the number and size distribution of the component axons of nerve V. The light microscopic image indicates one example of nerve V contains 2030 individual axons with a mean diameter of 1.4 μ; the electron image indicates the nerve contains 5794 axons with a mean diameter of 0.56 μ. The disparity between the two types of data is due to the large number of very small axons (0.2 μ or less in diameter) that course through the femoral nerve.     

贯众属的叶表皮特征

对贯众属（Cyrtomium）19种植物和近缘类群的15种植物的叶表皮形态特征进行了光学显微镜观察,并对包括贯众属3个亚系的模式种在内的12个种进行了扫描电镜观察。结果显示贯众属的叶表皮细胞为多边形或不规则形,垂周壁近平直、弓形、浅波状、波状至深波状。贯众属的气孔器分布于叶片下表皮,有无规则型、横列型和极附型三种类型,其中无规则型是主要的气孔器类型。气孔器表面观为宽椭圆形,长椭圆形,稀为近圆形,气孔外拱盖内缘近平滑、浅波状至啮齿-浅波状。大多数种类叶片表面角质膜具条纹,并常有条状隆起,或具颗粒等附属物。目前研究未发现可作为邢公侠二系四亚系诊断特征的明显的叶解剖特征。     

应用光镜和电镜对病虾组织细胞病理变化的观察与分析

应用光镜和电子显微镜技术比较研究正常与发病的中国对虾7种组织细胞病理变化,结果显示,病毒侵染后,对虾组织病理变化主要集中在消化系统的肝胰腺、中肠、胃等组织。在光镜下,可见消化道内壁上皮组织广泛受损,细胞大量坏死或空泡化;电镜下可见主要的细胞器如线粒体嵴大量断裂、粗面内质网严重扩张、溶酶体增生,病变细胞内出现大量变性的膜性结构等。观察结果为弄清对虾病毒引起的病症及致病机理打下基础。     

Preparation of chromosome spreads for electron (TEM,SEM, STEM), light and confocal microscopy

In the past, ultrastructural studies on chromosome morphology have been carried out using light microscopy, scanning electron microscopy and transmission electron microscopy of whole mounted or sectioned samples. Until now, however, it has not been possible to use all of these techniques on the same specimen. In this paper we describe a specimen preparation method that allows one to study the same chromosomes by transmission, scanning-transmission and scanning electron microscopy, as well as by standard light microscopy and confocal microscopy. Chromosome plates are obtained on a carbon coated glass slide. The carbon film carrying the chromosomes is then transferred to electron microscopy grids, subjected to various treatments and observed. The results show a consistent morphological correspondence between the different methods. This method could be very useful and important because it makes possible a direct comparison between the various techniques used in chromosome studies such as banding, in situ hybridization, fluorescent probe localization, ultrastructural analysis, and colloidal gold cytochemical reactionsAbbreviations CLSM confocal laser scanning microscope - EM electron microscopy - kV kilovolt(s) - LM light microscope - SEM scanning electron microscope - STEM scanning-transmission electron microscope - TEM transmission electron microscope     

Electron microscopy of fine needle aspiration biopsies of mediastinal and paramediastinal lesions

The ultrastructural cytologic study of fine needle aspiration (FNA) biopsies from eight cases with mediastinal and paramediastinal lesions is reported. In these cases, electron microscopy (EM) was essential in cytologically determining the correct type of the cancer cells. The results in these cases suggest that portions of FNA biopsies from deep sites, where aspiration is difficult or requires computed tomographic scanning, should be routinely processed for plastic embedding, so that EM examination can be performed if the cells are undifferentiated, scanty or poorly preserved by light microscopic examination. The proper cytologic identification of the cell might, in fact, have a major bearing on the therapeutic choices and on the prognosis.     

Reflection contrast microscopy of ultrathin sections in immunocytochemical localization studies: a versatile technique bridging electron microscopy with light microscopy

Reflection contrast microscopy (RCM) of ultrathin sections was recently introduced as a sensitive technique for visualization with enhanced definition in immunogold histochemistry. Experience of using RCM as a major tool in immunocytochemical research in different fields is summarized, e.g. oncology, nephrology and embryology. The sensitive visualization of immunocytochemical labels, gold particles or peroxidase-diaminobenzidine deposits in or on ultrathin sections, by RCM instead of electron microscopy is demonstrated. RCM of ultrathin sections is an adequate light microscopical alternative for immunoelectron microscopy, since an overview of both label and tissue is obtained with a high image definition and high contrast of label. In the studies presented, RCM is shown to provide a better gradation in staining intensity and staining pattern than other light microscopical methods. Moreover, a precise localization of multiple labels is obtained with this method. Besides the applications shown, ultrathin section visualization by RCM is very useful for correlative light- and electron microscopical studies of fine structures. Commercially available fluorescence microscopes can be adapted for proper RCM functioning; an adaptation scheme and list of microscopes tested is provided.     

Near-infrared branding efficiently correlates light and electron microscopy

The correlation of light and electron microscopy of complex tissues remains a major challenge. Here we report near-infrared branding (NIRB), which facilitates such correlation by using a pulsed, near-infrared laser to create defined fiducial marks in three dimensions in fixed tissue. As these marks are fluorescent and can be photo-oxidized to generate electron contrast, they can guide re-identification of previously imaged structures as small as dendritic spines by electron microscopy.     

Binding of wheat germ agglutinin in the matrix of rat tracheal cartilage

Summary The binding of wheat germ agglutinin (WGA) to the extracellular matrix of rat tracheal cartilage was studied at both the light and electron microscopic levels. The detection of binding sites was accomplished by a postembedment method using the direct fluorescence technique for light microscopy and the avidin—biotin bridging system for electron microscopy. Distinct fluorescence was observed in the pericellular region of chondrocytes, and this fluorescence was completely removed after treatment with 4 M guanidine hydrochloride. By electron microscopy, the reaction products as found in the pericellular region were not observed in the interterritorial collagenous matrix, confirming a similar distribution as found by fluorescence microscopy. These results show that WGA-binding sites are present in pericellular matrical substances, which are known to be rich in proteoglycan and glycosaminoglycan complexes and which exhibit similar staining with antibodies to proteoglycans or with cationic dyes. As WGA binds toN-acetyl-glucosamine andN-acetyl-neuraminic acid residues, the pericellular matrix of rat tracheal cartilage appears to consist of proteoglycan having a high concentration of these saccharides.     

Morphological and functional aspects of volatile-producing glands in bees (Hymenoptera: Apidae)

Abstract In this paper we focus on the occurrence and morphological aspects of exocrine glands in several bee species. Morphology of head labial, mandibular, Dufour, and abdominal tegumentar glands was investigated under light microscopy, scanning electron microscopy and transmission electron microscopy. Most of such glands present cells with cytoplasm homogeneous and acidophilic, or contain small apparently empty vacuoles. The cytoplasm cells' ultrastructure showed a well developed smooth endoplasmic reticulum, many polymorphic mitochondria, rare Golgi, lipid droplets, myelin figures, and many basal and apical plasma membrane infoldings. All these results are discussed in the text.     

High data output and automated 3D correlative light-electron microscopy method

Correlative light/electron microscopy (CLEM) allows the simultaneous observation of a given subcellular structure by fluorescence light microscopy (FLM) and electron microscopy. The use of this approach is becoming increasingly frequent in cell biology. In this study, we report on a new high data output CLEM method based on the use of cryosections. We successfully applied the method to analyze the structure of rough and smooth Russell bodies used as model systems. The major advantages of our method are (i) the possibility to correlate several hundreds of events at the same time, (ii) the possibility to perform three-dimensional (3D) correlation, (iii) the possibility to immunolabel both endogenous and recombinantly expressed proteins at the same time and (iv) the possibility to combine the high data analysis capability of FLM with the high precision-accuracy of transmission electron microscopy in a CLEM hybrid morphometry analysis. We have identified and optimized critical steps in sample preparation, defined routines for sample analysis and retracing of regions of interest, developed software for semi/fully automatic 3D reconstruction and defined preliminary conditions for an hybrid light/electron microscopy morphometry approach.     

Fine structure of the parotid gland in the crest-tailed marsupial-rat (Dasyuroides byrnei).

The parotid gland of Dasyuroides byrnei was examined by light microscopy, and transmission and scanning electron microscopy. The acini were composed predominantly of seromucous cells with a few mucous cells. The seromucous cells were light or dark cells containing acidophilic spherical granules of moderate to high electron density and had well-developed cytoplasmic organelles-ordinary mitochondria and large mitochondria with tubular cristae, RER with vesicular or tubular elements, and Golgi apparatus with lamellae, vesicles and vacuoles. The mucous cells had basophilic amorphous granules of low electron density, like those of ordinary mucous cells. The intercalated ducts were composed of simple cuboidal light cells having a few electron-dense granules. The striated ducts consisted of tall columnar light cells containing numerous vesicles and mitochondria with tubular cristae, the same as found in acinar seromucous cells.     

Effects of the cationic protein poly-L-arginine on airway epithelial cells in vitro

BACKGROUND: Allergic asthma is associated with an increased number of eosinophils in the airway wall. Eosinophils secrete cationic proteins, particularly major basic protein (MBP). AIM: To investigate the effect of synthetic cationic polypeptides such as poly-L-arginine, which can mimic the effect of MBP, on airway epithelial cells. METHODS: Cultured airway epithelial cells were exposed to poly-L-arginine, and effects were determined by light and electron microscopy. RESULTS: Poly-L-arginine induced apoptosis and necrosis. Transmission electron microscopy showed mitochondrial damage and changes in the nucleus. The tight junctions were damaged, as evidenced by penetration of lanthanum. Scanning electron microscopy showed a damaged cell membrane with many pores. Microanalysis showed a significant decrease in the cellular content of magnesium, phosphorus, sodium, potassium and chlorine, and an increase in calcium. Plakoglobin immunoreactivity in the cell membrane was decreased, indicating a decrease in the number of desmosomes CONCLUSIONS: The results point to poly-L-arginine induced membrane damage, resulting in increased permeability, loss of cell-cell contacts and generalized cell damage.     

Sex chromosomes at early meiosis in three wood mice species of the genus Apodemus (Rodentia, Muridae)

The prophase of the first meiotic division was studied in field mice of the species Apodemus (Sylvaemus) flavicollis, A. (S.) ponticus, and A. (S.) uralensis by light and electron microscopy. The karyotypes of the species were described on the base of electron microscopy of synaptonemal complexes in spermatocytes I. The axial elements of the sex chromosomes at early-middle pachytene can synapse along the major portion of the Y axis; at late pachytene-early diplotene, the synapsis region shrinks; and at diakinesis-metaphase I, X and Y chromosomes associate tail-to-tail in all species studied. The behavior of sex chromosomes in the synapsis in the species studied was quite uniform. The results are discussed in the context of earlier data on the behavior of sex chromosomes in various rodent species in meiosis prophase I and their banding.     

Comparison of Light and Electron Microscopic Determinations of the Number of Bacteria and Algae in Lake Water

Determinations of the number of microorganisms in lake water samples with the bright-field light microscope were performed using conventional counting chambers. Determinations with the fluorescence microscope were carried out after staining the organisms with acridine orange and filtering them onto Nuclepore filters. For transmission electron microscopy, a water sample was concentrated by centrifugation. The pellet was solidifed in agar, fixed, dehydrated, embedded in Epon, and cut into thin sections. The number and area of organism profiles per unit area of the sections were determined. The number of organisms per unit volume of the pellet was then calculated using stereological formulae. The corresponding number in the lake water was obtained from the ratio of volume of solidified pellet/volume of water sample. Control experiments with pure cultures of bacteria and algae showed good agreement between light and electron microscopic counts. This was also true for most lake water samples, but the electron microscopic preparations from some samples contained small vibrio-like bodies and ill-defined structures that made a precise comparison more difficult. Bacteria and small blue-green and green algae could not always be differentiated with the light microscope, but this was easily done by electron microscopy. Our results show that transmission electron microscopy can be used for checking light microscopic counts of microorganisms in lake water.     

Plasmolysis of Pteridium protoplasts: A study using light and scanning-electron microscopy

A study was undertaken using gametophytes of the fern Pteridium aquilinum to examine the effects of plasmolysis on the topography of protoplasts. Methods are described whereby the surfaces of non-isolated protoplasts can be observed in the plasmolysed condition using scanning electron microscopy. Plasmolysed gametophytes were also examined in the light microscope using differential interference contrast and ultra-violet fluorescence microscopy after staining with fluorescein diacetate. With scanning electron microscopy, plasmolysed protoplast surfaces appeared smooth with no evidence of wrinkling or infolding of excess membrane. The formation of irregular-shaped protoplasts, protoplasmic threads, subprotoplasts, and protoplasmic networks covering internal wall surfaces all provided evidence for strong wall adhesion of the protoplasm. The availability of membrane for uptake into folds or vesicles is therefore thought to be minimal. Transmission electron microscopy showed some protoplasmic threads to be plasmodesmata, the remainder being cell-wall contact points. Remnants of these threads were occasionally observed on isolated protoplasts in both the light and electron microscopes.     

Testicular chromosomes of Gallus domesticus

Summary Testes of Gallus domesticus were studied (a) by light microscopy after hypotonic treatment followed by acetic-alcohol fixation and airdrying and (b) by electron microscopy of osmium-fixed, araldite-embedded material, some of which was pretreated with hypotonic solutions.The following conclusions were reached: (i) The number of chromosome pairs at meiosis is constant and is most probably 40 (although 39 or 38 is possible). (ii) The diploid chromosome number at mitotic metaphase cannot be certainly determined by light microscopy but there is no reason to suppose it is not double the number of meiotic bivalents. (iii) No essential difference in structure was found between long and short bivalents at meiosis by light or electron microscopy; the lengths of the bivalents at pachytene form a continuous series, (iv) Some short bivalents appear to contain less material per unit length than long ones; this could explain why these chromosomes cannot always be resolved by light microscopy when fully contracted. (v) So-called macro- and micro-chromosomes differ only in size, but not in behaviour, at mitosis and meiosis.     

Ultrastructure in cell biology: do we still need it?

John Heuser is being honored in this special issue for his enormous contributions to cell biology using morphological approaches. Foremost in this context is his ability to use light and electron microscopy to visualize structures and processes such that the information has both scientific and artistic value. The beauty of his images helps to focus the observer more intensely on the scientific messages, which have been numerous and important. His recent studies of living cells using state-of-the-art light and video microscopy fits into a general pattern of a huge explosion in the application of these methods worldwide that is revolutionizing cell biology. However, whereas John Heuser continues to use light microscopy (LM) for a low-resolution global and dynamical overview he then moves on to the electron microscopy (EM) level to see the details; in this he is--unfortunately--in a minority; and EM is an approach that a majority of today's cell biologists never use. The continued drop in EM usage has already been articulated in recent reviews. Here, I suggest that an additional problem for EM in cell biology, in its continued crises, is the declining number of scientists who can confidently interpret the--admittedly--complex information in most electron micrographs of cells. A major re-education is needed, or cell biology as a discipline will have a real problem in the 21st century.     

Structural and Ultrastructural Characteristics of Bone-Tendon Junction of the Calcaneal Tendon of Adult and Elderly Wistar Rats

Tendons are transition tissues that transfer the contractile forces generated by the muscles to the bones, allowing movement. The region where the tendon attaches to the bone is called bone-tendon junction or enthesis and may be classified as fibrous or fibrocartilaginous. This study aims to analyze the collagen fibers and the cells present in the bone-tendon junction using light microscopy and ultrastructural techniques as scanning electron microscopy and transmission electron microscopy. Forty male Wistar rats were used in the experiment, being 20 adult rats at 4 months-old and 20 elderly rats at 20 months-old. The hind limbs of the rats were removed, dissected and prepared to light microscopy, transmission electron microscopy and scanning electron microscopy. The aging process showed changes in the collagen fibrils, with a predominance of type III fibers in the elderly group, in addition to a decrease in the amount of the fibrocartilage cells, fewer and shorter cytoplasmic processes and a decreased synthetic capacity due to degradation of the organelles involved in synthesis.     

Immunocytochemical localization of cathepsin H in rat kidney. Light and electron microscopic study

Light and electron microscopic localization of cathepsin H in rat kidney was studied using post-embedding immunocytochemical techniques. For light microscopy, Epon sections of the kidney were stained by immunoenzyme method after removal of Epon and for electron microscopy, ultrathin sections of the Lowicryl K4M-embedded material were labeled by protein A-gold (pAg) technique. By light microscopy, fine granular staining was found in throughout the nephron, but the staining intensity considerably varied. The strongest staining was noted in the S1 segment of the proximal tubules followed by the S2 and S3 segments and the medullary collecting tubules. The glomeruli, the distal tubules, and the cortical collecting tubules were weakly stained. By electron microscopy, a gold label was found exclusively in lysosomes, which showed various sizes and labeling intensity. The results were quite consistent with the light microscopic results. The labeling intensity tended to increase as the matrix of lysosomes was condensed. Quantitative analysis of the labeling density of lysosomes demonstrated that the highest labeling density is found in the S1 segment of the proximal tubules and the labeling density of other renal segments is significantly low levels. The results indicate that a main site for cathepsin H in rat kidney is the S1 segment of the proximal tubules.     

Light and electron microscopic demonstration of sialic acid residues with the lectin from Limax flavus: a cytochemical affinity technique with the use of fetuin-gold complexes

The development of a cytochemical affinity technique for the demonstration of sialic acid residues by light and electron microscopy is reported. The lectin from the slug Limax flavus, with its narrow specificity for N-acetyl- and N-glycolylneuraminic acid, was applied to tissue sections. Subsequently fetuin-gold complexes were used to visualize the tissue-bound lectin. Different cytochemical controls, including sugar inhibition tests, neuraminidase digestion, the use of fetuin-gold complexes alone, or acid hydrolysis of sections, proved the specificity of the technique. Postembedding staining was performed on frozen, paraffin, or semithin resin sections for light microscopy and on thin sections from low temperature Lowicryl K4M-embedded material for electron microscopy. The distribution of sialic acid residues in rat pancreas, liver, and colonic mucosa was investigated.