Study of the root nodules in some species of the Papilionaceae subfamily by scanning electron microscopy

Nitrogen-fixing nodules from 16 species in 6 tribes of the sub-family Papilionaceae have been examined by scanning electron microscopy. The structure of infection threads was similar in all the studied papilionoid species except Lupinus polyphillus. In this species the infection threads were found in young nodules only. The morphology of bacterioids and the character of their "package" are determined by the host plant genotype. The obtained results are discussed in relation to the evolution of the legumes.     

Cell fine structures observed by scanning electron microscopy]

The scanning electron microscope (SEM) provides vivid seemingly three dimensional images which are easier to understand for us than transmission electron microscopic images. For this point of view scanning electron microscopy is advantageous in morphological researches of cell fine structures. Nevertheless, there were few studies in this field, because SEM had much lower resolution than transmission electron microscope (TEM) and because there was no adequate method to reveal intracellular structures. In recent years, however, the resolution of SEM has been markedly improved and the specimen preparation techniques have also advanced. In this paper, some of our preparation technique for revealing cell surface structures or intracellular structures, in particular, osmium-DMSO-osmium method, and the results observed by these methods were described. 1) Nucleus. The nucleus was wrapped with a nuclear envelope that consisted of two membranes enclosing a narrow space. On the surface of the envelope many nuclear pores were observed. 2) Endoplasmic reticulum (ER). Rough ER consisted of flattened cisternae, arranged in parallel. The surface were studded with many ribosomes which were often arranged spirally to form polysomes. Smooth ER consisted of tubules. 3) Golgi complex. a) The Golgi stacks were all linked by anastomosing. b) Connection between Golgi stacks and rough ER was often observed. c) Cisternae in a Golgi stack were connected each other. 4) Mitochondria. The mitochondrion was bounded by 2 sheets of unit membrane and the inner membrane projected into the interior of the organelles to make mitochondrial cristae.     

小鼠生精细胞染色质与染色体构筑的扫描电镜研究（简报）

It remains unclear about the intermediate construction of chromosome due to its highly compact nature and the limitation in methods. The present study was designed to investigate the construction of chromatin and mitotic chromosome in situ with scanning electron microscopy. Mouse testes were selected as the material, because of in which the spermatogenic cells divide actively and successively to form the sperm. Such a feature would be able to study the structure of mammalian chromatin and chromosomes along with the change of nuclear cycle. The animal were perfused with 200 ml of 0.075 mol/L KCl hypotonic solution to remove blood and placed for 15-20 min on ice followed by 0.5% glutaraldehyde and 0.5% formaldehyde for fixing. Through treated by the routine process of fractured and freeze dried with t-butyl alcohol, the specimens were then coated with a 3 nm thick platinum and observed with Hitachi S-430 scanning electron microscopy. It was found that the hypotonic treatment with 0.075 mol/L KCl solution was suit for demonstrating the nuclear structure, when the organelles were well preserved. The chromatin fibers of 10-30 nm and 80-125 nm in diameter could be recognized in the interphase nuclei, which were arranged losely at the region of euchromatin, and folded with each other into chromatin masses at the region of heterochromatin, while the chromatin fibers with the diameter of 80-125 nm often could be viewed on the mitotic chromosomes. Since its presence in interphase nuclei and mitotic chromosomes, it was considered that the chromatin fibers with 80-125 nm in diameter might play a role in the condensation of chromosome, serve as a type of the intermediate structure.     

Study of the surface of cytolytic T-lymphocytes by scanning electron microscopy

Scanning electron microscopy revealed three types of cytolytic T lymphocytes (CTL) interacting with target cells (TC). The type I cells occurred as smooth spherical lymphocytes with single microvilli; the type II rounded or oval cells were uniformly covered with microvilli; the type III cells were marked by an irregular shape, were densely covered with microvilli, while their surface was folded or tuberous. Within the first several minutes after absorption the surface of TC was largely covered by the type I lymphocytes. The proportion of the type III cells rose with the time of interaction. At the beginning it was 8-9%, reaching 30-71% after 30-60 minutes of incubation. It is assumed that the 3 types of the cells described mirror 3 stages of CTL activation. The increase in number of microvilli and appearance of the membranous folds may be a consequence of exocytosis and incorporation of the membrane of secretory granules into the plasma membrane of lymphocytes. The data obtained support the authors' assumption about the secretory mechanism of the action of CTL, whose contact with TC stimulated secretion activation.     

Birefringent objects visualized by circular polarization microscopy

In the standard polarizing microscope, birefringent material appears bright only if its optic axis is oblique to the axes of the polarizer and analyzer filters; consequently, an object may be visualized as several disconnected bright regions. This confusing appearance is avoided if the crossed plane polarizers of the conventional microscope are replaced by circular polarizers of opposite handedness. All orientations of the optic axis in the focal plane then become equivalent; objects generally appear uniformly bright. Ordinary microscopes are easily modified to use this technic with readily available components.     

Method of preparing suspended cells for scanning electron microscopy]

To preserve the lifetime morphology of the surface of suspended cells, these must be fixed in suspensions. The subsequent stages of cell preparation for scanning electron microscopy (dehydratation, critical point drying, coating) are considerably facilitated if fixed cells are preliminary attached to some substrate surface. An effective substrate should provide a firm rather than selective attachment of the overwhelming majority of fixed cells; the substrate should be also available for a wide application. The trial of different types of substrates showed a sufficient effectivity of plates made of commercial aluminium foil. In tests with murine embryonal and transformed fibroblasts as well as with human blood leukocytes, in average 90 per cent of cells fixed with glutaraldehyde in suspensions got attached to foil substrate surfaces; the fixed cells both settled from suspension and attached were seen distributed evenly on the substrate surface. The use of aluminium foil substrates made it possible to study the surface topography of some types of suspended cells.     

Adaptation of the tetrazolium method for testing the seed viability, and scanning electron microscopy study of some Western European orchids

A modified tetrazolium method was formulated for use with seeds of Western European orchids. The sequence of treatments which gave the highest percentage of coloured (i.e. viable) embryos was: (1) pretreatment in a solution of 5% (w/v) Ca(OCl)₂+ 1% (v/v) Tween-80, (2) soaking for 1 day in sterile water, (3) the classical tetrazolium test. The optimal duration of the pretreatment in Ca(OCl)₂+ Tween-80 depends upon the species, and to investigate the effect a scanning electron microscopy study was performed on the testa of 3 species. For a given species, the optimal pretreatment period was not affected by the year of harvest or the source of the seed lots.     

Study of the microcirculation of the wall of the stomach of white rats by the vital microscopy method]

Microcirculation of the gastric wall in the albino rat was studied by the method of vital microscopy in passing light. The conditions of the experiment permitted watching the microcirculation state during 5-6 hours. Intravital morphometry of the microcirculation bed vessels was made. The obtained data concerning the intravital angioarchitectonics of the gastric wall vascular bed were in good agreement with a conventional scheme of its blood supply. The role of arteriolevenular anastomoses in redistribution of blood within the bloodflow pathways was noted.     

Subendocardial vascular plexus of the human ventricular myocardium in ontogenesis studied by scanning electron microscopy]

By means of scanning electron microscopy of native and corrosive preparations peculiarities of endocardium of the papillary-trabecular apparatus of the human heart ventricle and its vessels have been studied in ontogenesis. Folds of the endocardium increase its contact area with blood and also increase reserve of its elasticity at diastole. The subendocardial vascular plexus is formed by capillary-like venous sinusoids. They situate in the direction to the compact myocardium at the depth of 300-500 mcm. They are supposed to perform resorptive functions of nutritive substances and oxygen from blood, which is in the ventricular cavities and to smooth blood pressure between blood in the ventricular cavities and vessels of the compact myocardium.     

Study of the lympho- and hemomicrocirculatory bed of lymph nodes by scanning electron microscopy

The spatial interrelation of the lymphatic and blood beds has been studied in the lymph node together with its stromal and lymphoid elements, using scanning electron microscopy of corrosive casts and native preparations. There is a great variety of pathways for lymph transport and blood vessels in dependence of their localization in the lymph node. Of a special interest are the pathways of lymphocytes migration across the walls in the cortical and medullary sinuses and in the postcapillary venules, as well as participation of the reticular tissue in the lymph node immune reactions.     

A method for obtaining histological sections from tissue previously studied by scanning electron microscopy

An accelerated method of paraffin embedding of tissue specimens previously examined with scanning electron microscopy is proposed aimed to obtain sections for routine histological examination. The tissue is passed through acetone, absolute alcohol, alcoholic-oil celloidin solution, chloroform to be eventually mounted into paraffin. The method allows obtaining good quality sections within 24 hours.     

Electromechanical imaging of biomaterials by scanning probe microscopy

The majority of calcified and connective tissues possess complex hierarchical structure spanning the length scales from nanometers to millimeters. Understanding the biological functionality of these materials requires reliable methods for structural imaging on the nanoscale. Here, we demonstrate an approach for electromechanical imaging of the structure of biological samples on the length scales from tens of microns to nanometers using piezoresponse force microscopy (PFM), which utilizes the intrinsic piezoelectricity of biopolymers such as proteins and polysaccharides as the basis for high-resolution imaging. Nanostructural imaging of a variety of protein-based materials, including tooth, antler, and cartilage, is demonstrated. Visualization of protein fibrils with sub-10nm spatial resolution in a human tooth is achieved. Given the near-ubiquitous presence of piezoelectricity in biological systems, PFM is suggested as a versatile tool for micro- and nanostructural imaging in both connective and calcified tissues.     

Structure of immunoglobulin G by scanning tunnelling microscopy

Scanning tunnelling microscopy (STM) has been used to examine the shape of individual immunoglobulin G (IgG) molecules deposited onto a graphite surface. IgG was chosen for this study as it has a well-characterized and distinctive three-dimensional structure. The micrographs clearly reveal the IgG molecule as trilobed, corresponding with the known structural organization of IgG. Comparison of these images with the structure of IgG determined by X-ray crystallography shows that the STM images are consistent with the crystal structure. This illustrates that STM is a valuable technique for examining protein structure, allowing rapid determination of the overall molecular shape that is consistent with more established techniques.