Transmission electron microscopy (TEM) provides high resolution images, which are useful in studying ultrastructure of cells and tissues. We have to use very thin section about 60~100 nm thickness due to poor penetration power of the conventional TEM at 100 kV. To overcome this limitation, TEMs using higher accelerating voltage have been developed. TEMs can be categorized into conventional TEM, intermediate TEM, high voltage TEM (HVEM), and ultrahigh voltage TEM according to their accelerating voltage. HVEM using 500~1,000 kV has an enough penetration power to observe thick specimen up to 3~4 micro, which is useful understanding 3 dimensional configuration of the cell and tissue. HVEM was built up in Korea Basic Science Institute (KBSI, Daejeon, Korea) at 2004, maximum accelerating voltage is 1.3 MV in Korea. Many results showed up to the present various fields of science such as medical science, biology, agriculture and so on. Here, we briefly summarize recent biomedical applications of HVEM to provide an insight of HVEM for morphologist.
Agriculture ; Biology ; Electrons ; Korea ; Microscopy, Electron ; Microscopy, Electron, Transmission

Objective To establish an efficient method for isolating migrasomes from RAW264.7 macrophages and identifying these isolated migrasomes. Methods Scanning electron microscopy was used to observe the morphological characteristics of migrasomes produced by RAW264.7 cells. A 0.45 μm filter was employed for reverse filtration and elution to isolate the migrasomes. The morphological characteristics of the migrasomes were then observed using transmission electron microscopy. Western blot analysis was performed to determine the expression of characteristic markers of the migrasomes. The RNA carried by the migrasomes was analysed by using LabChip bioanalyzer. Results Scanning electron microscopy revealed that the migrasomes, with membranous structures, were attached to the tip or bifurcation of the retraction fiber formed in the tail of RAW264.7 cells. Transmission electron microscopy showed that the isolated migrasomes had a typical oval vesicle-like structure with wrinkled membrane surfaces. Western blot analysis confirmed the expression of the characteristic markers phosphatidylinositol glycan anchor biosynthesis class K (PIGK), epidermal growth factor domain-specific O-linked N-acetylglucosamine transferase (EOGT) and tetraspanin 4 (TSPAN4) in the migrasomes, while the EV (extracellular vesicle) markers tumor susceptibility gene 101 (TSG101) and Arabidopsis homolog of apoptosis-linked gene 2-interacting protein X (ALIX) were not detected. Furthermore, the isolated migrasomes were found to be rich in small RNA, which were approximately 25-200 nt in length. Conclusion A method for the extraction of well-structured and high quality migrasomes from macrophages is established.
Extracellular Vesicles ; Microscopy, Electron, Transmission ; RNA ; Macrophages

OBJECTIVE: We performed this study to evaluate the effects of Sil-Select and Percoll in sperm preparation. METHODS: Semen samples of 22 patients with normal sperm parameters by WHO criteria were divided into two equal parts and prepared with Percoll and Sil-Select. After completion of semen preparation procedures with Percoll and Sil-Select, sperm concentration, motility and morphology using strict criteria were evaluated in each group and all semen samples were fixed and stained for transmission electron microscopy(TEM). RESULTS: There were no significant diffrences in sperm concentration, percentage of motile spermatozoa and percentage of normal spermatozoa in morphology evaluation using strict criteria under light microscopy between Percoll and Sil-Select-treated groups. However, the percentage of normal shape and position of acrosome, and normal helix assembly of mitochondria under TEM were significantly higher in the Sil-Select-treated group compared to Percoll-treated group (p < 0.001, p < 0.001, p < 0.001). CONCLUSIONS: These data demonstrate that Sil-Select is less detrimental to the acrosome and mitochondria of spermatozoa in sperm preparation compared to Percoll.
Acrosome ; Humans ; Male ; Microscopy ; Microscopy, Electron, Transmission ; Mitochondria ; Semen ; Spermatozoa*

Morphological characteristics of hyphal interaction between Grifola umbellata (Pers. Ex Fr.) Pilat and its companion fungus which related to sclerotia formation from hyphae were investigated by external observations, light microscopy and transmission electron microscopy (TEM). External observations showed that a dense antagonism line was formed by both G. umbellata and companion fungus after their hyphae contacted each other in dual culture. Many hyphal strands emerged on the colony of G. umbellata and differentiated to sclerotia from where hyphal strands crossed. Light microscope observations revealed the process of antagonism line formation. Mature antagonism with structural differentiation, was composed of three main layers: the rind, the rind underlayer and the hypha layer. TEM observations showed that after colonies hyphal contact, a series of reactions always occurred in both G. umbellata and companion fungus. Cells in the center of antagonism line were dead. Cells of G. umbellata adjacent to the antagonism line were usually large and hollow, with unilateral thickened wall, whereas those of companion fungus were empty, with thin or thick wall. Both hyphal interaction at the antagonism line may be one of the main reasons for sclerotia of G. umbellata differentiation from hypha.
Friends* ; Fungi* ; Grifola* ; Humans ; Hyphae ; Microscopy ; Microscopy, Electron, Transmission

PURPOSE: This study was carried out to clarify the histological characteristics of the interface of the corneal stroma and Descemet's membrane of the human eye. METHODS: Nighteen donor eyes without corneal pathology were examined by scanning and transmission electron microscopy. The Descemet's membrane including the corneal endothelium was cheked for scanning electron microscopy. The junctional characteristics of the posterior corneal stroma and Descemet's membrane was examined by transmission electron microscopy. RESULTS: The scanning electron microscopy showed that collagen sheet faced each other at the right angle near the Descemet's membrane and penetrated the Descemet's membrane with the irregular arrangement. The transmission electron microscopy showed that the electron-dense collagen filaments extended to the posterior stroma from Descemet's membrane. The arrangement of electron-dense collagen filaments paralleled with the arrangement of the collagen fibrils of the posterior stroma. CONCLUSIONS: The interface of the corneal stroma and Descemet's membrane was composed of two-typed extracellular materials without the intercellular specificatons.
Collagen ; Corneal Stroma* ; Descemet Membrane* ; Endothelium, Corneal ; Humans ; Microscopy, Electron ; Microscopy, Electron, Scanning ; Microscopy, Electron, Transmission ; Pathology ; Tissue Donors

PURPOSE: To investigate of the histological characteristics of the lattice degeneration of the human peripheral retina. METHODS: The histological characteristics of the lattice degeneration of the retina was checked by flat preparation and serial section of the lattice lesion in three eyes was investigated by transmission electron microscopy. RESULTS: Flat preparation showed lattice lesion with a hole at the lateral margin with overlying sclerotic vessel and pigment clumping within the lesion. The ultrastructural initial findings showed that the collagen filament in the vitreous cavity was continuous with Muller fiber of the retina with the defect of the inner retina. The full-thikness defect of the sensory retina leaded to the retinal hole. The vascular wall was replaced and occluded by fine fibrillar collagen. The glial cell proliferated into the neural tissue of the sensory retina. These glial cells may secrete long spacing collagen (LSC) and curvilinear material shown at the area of the sensory retinal defect and near the vitreoretinal interface. CONCLUSIONS: These results suggest that the thinning of the retina occurs from the inner retina leading to retinal hole as the lattice degeneration progresses. LSC and curvilinear material are suggestive of derivatives derived from the extracellular material secreted from the glial cell.
Collagen ; Fibrillar Collagens ; Humans ; Microscopy, Electron ; Microscopy, Electron, Transmission ; Neuroglia ; Retina* ; Retinal Perforations ; Retinaldehyde*

Lung fluke, Paragonimus heterotremus, is a flatworm causing pulmonary paragonimiasis in cats, dogs, and humans in Southeast Asia. We examined the ultrastructure of the testis of adult P. heterotremus with special attention to spermatogenesis and spermiogenesis using scanning and transmission electron microscopy. The full sequence of spermatogenesis and spermiogenesis, from the capsular basal lamina to the luminal surface, was demonstrated. The sequence comprises spermatogonia, spermatocytes with obvious nuclear synaptonemal complexes, spermatids, and eventual spermatozoa. Moreover, full steps of spermatid differentiation were shown which consisted of 1) early stage, 2) differentiation stage representing the flagella, intercentriolar body, basal body, striated rootlets, and electron dense nucleus of thread-like lamellar configuration, and 3) growing spermatid flagella. Detailed ultrastructure of 2 different types of spermatozoa was also shown in this study.
Animals ; Male ; Microscopy, Electron, Scanning ; Microscopy, Electron, Transmission ; Paragonimus/*physiology/*ultrastructure ; Spermatogenesis ; Spermatozoa/ultrastructure ; Testis/ultrastructure

PURPOSE: To evaluate 20% ethanol toxicity on the rabbit corneal epithelium, ethanol-treated rabbit corneas were examined with electron microscopy. METHODS: Rabbit corneas(24 eyes) were treated with 20% ethanol for 30 seconds, 1 minute, and 2 minutes by using LASEK(Laser Assisted Subepithelial Keratomileusis) instruments, and washed with sterile water. Zero time, 1, 3, 5 days after ethanol treatment, corneas were excised and examined with scanning and transmission electron microscopy. RESULTS: Widespread damage or disappearance of microvilli and local breaks of intercellular junction were observed. The changes were more severe in corneas with longer ethanol treament. In corneas with over 1 minute ethanol treatment, slough of superficial corneal epithelium was shown and increased with time. It was difficult to recognize microvilli or distinctive intercellular junction in corneas with 2 minute-treament. These pathologic changes persisted 5 days after ethanol-treatment. CONCLUSIONS: From these results, 30 seconds to 1 minute-ethanol treatment is recommended in corneal surgery to avoid severe, persisting damage of superficial corneal epithelium.
Cornea ; Epithelial Cells* ; Epithelium, Corneal ; Ethanol ; Intercellular Junctions ; Microscopy, Electron ; Microscopy, Electron, Transmission ; Microvilli ; Water

BACKGROUNDTrichosporon asahii (T. asahii) is one of the most important pathogenic fungus in the genus of trichosporon. Although the species identification of T. asahii was based upon the complicated results of morphologic, biochemical and biologic examination, the morphology characteristic is still the first clue to the species. Some common structures of T. asahii had been described such as arthrofilaments and arthroconidia, but other important structures of T. asahii were unclear.

METHODSSix strains of T. asahii were incubated on the slant and micro culture of Sabouraud's dextrose agar at 30 degrees C for 7 days. Samples were fixed using 2% paraformaldehyde and 2.5% glutaraldehyde. T. asahii was observed under scanning electron microscope and transmission electron microscope.

RESULTSThe detailed characteristics of the diverse sites of germination, as well as some uncommon structures such as giant cell, sarcinate, and club-shaped macroconidia, were presented. The pseudohyphae of T. asahii were noted to produce true hyphae, either along the longitude axis or on the flank. T. asahii was noted to have blastic and thallic conidiation. Digitated branches, trichoid structures and septa inside the spores were detected.

CONCLUSIONThese results may add our knowledge to the structure and development of T. asahii.

OBJECTIVETo design a kind of biomimetic polypeptide of dentin matrix protein-1 (DMP-1), which can bind to dentine collagen fibers and initiate mineralization.

METHODSA novel polypeptide was developed by connecting the collagen binding domain of DMP-1 "DSESSEEDR" to the hydrophilic C-terminal of amelogenin "TKREEVD". The polypeptide was synthetically prepared by standard solid-phase peptide synthesis. Human dentine slices were completely demineralized by hydrochloric acid to expose the dentine collagen. Fluorescein isothiocyanate coupled polypeptide was applied to the exposed dentine collagen. Fluorescent microscopy was used to examine the polypeptide specially bond to the dentine collagen. Nucleation and growth of hydroxyapatite was initiated by immersing the polypeptide into calcium chloride and sodium hypophosphate solutions respectively. Scanning electron microscopy (SEM), transmission electron microscopy (TEM) and selected area diffraction (SAD) were used to examine the hydroxyapatites formed. RESULTS Fluorescent dentine collagen was identified in the demineralized dentine specimens. Nucleation and growth of crystals were detected after immersing the polypeptide into calcium chloride and sodium hypophosphate solutions by SEM and TEM. SAD confirmed the crystals were hydroxyapatites.

CONCLUSIONThe polypeptide of "DSESSEEDRTKREEVD" can simulate DMP-1 binding collagen and initiate hydroxyapatite nucleation and growth. It may be a potential molecular tool for dentine remineralization.