Transmission electron microscopy (TEM) of equine conceptuses at 14 and 16 days of gestation

2010 ◽  
Vol 22 (2) ◽  
pp. 405 ◽  
Author(s):  
Ingrid Walter ◽  
Waltraud Tschulenk ◽  
Sven Budik ◽  
Christine Aurich
Keyword(s):  
Electron Microscopy ◽  
Transmission Electron Microscopy ◽  
Cell Surface ◽  
Specific Cell ◽  
Fluid Uptake ◽  
Embryonic Disc ◽  
Transmission Electron ◽  
Cellular Junctions ◽  
Endodermal Cells ◽  
Specific Cell Surface

The present study gives a detailed ultrastructural description of equine conceptuses at Day 14 (n = 2) and Day 16 (n = 3) after ovulation. Whereas on Day 14 only primitive structures were seen, on Day 16 neurulation and formation of mesodermal somites had taken place. The ectoderm of the embryo itself and the surrounding trophoblast ectodermal cells were characterised by specific cell surface differentiations. At the embryonic ectodermal cell surface (14 and 16 days) remarkable protruded and fused cytoplasmic projections were seen, typically associated with macropinocytotic events involved in macromolecule and fluid uptake. This finding adds an important point to the expansion mode of the hypotone equine conceptus, which is characterised by ‘uphill’ fluid uptake. Numerous microvilli and coated endocytotic pits at the apical trophoblast membrane emphasised its absorptive character. Endodermal cells were arranged loosely with only apically located cellular junctions leaving large intercellular compartments. At the border of the embryonic disc apoptotic cells were regularly observed indicating high remodelling activities in this area. Conspicuous blister-like structures between ectoderm and mesoderm were seen in the trilaminar part of Day-14 and -16 conceptuses. These were strictly circumscribed despite not being sealed by cellular junctions between germinal layers. It is possible that these blisters are involved in embryo positioning; however, further studies are needed to verify this.

scholarly journals Same Serial Section Correlative Light and Energy-filtered Transmission Electron Microscopy

2003 ◽  
Vol 51 (5) ◽  
pp. 605-612 ◽  
Author(s):  
Ying Ren ◽  
Michael J. Kruhlak ◽  
David P. Bazett-Jones
Keyword(s):  
Electron Microscopy ◽  
Transmission Electron Microscopy ◽  
Light Microscope ◽  
High Rate ◽  
Specific Cell ◽  
3D Analysis ◽  
Serial Sections ◽  
Correlative Imaging ◽  
Transmission Electron ◽  
Rate Of Success

Correlative imaging of a specific cell with both the light microscope and the electron microscope has proved to be a difficult task, requiring enormous amounts of patience and technical skill. We describe a technique with a high rate of success, which can be used to identify a particular cell in the light microscope and then to embed and thin-section it for electron microscopy. The technique also includes a method to obtain many uninterrupted, thin serial sections for imaging by conventional or energy-filtered transmission electron microscopy, to obtain images for 3D analysis of detail at the suborganelle level.

The translaminal fibrils of the human amnion basement membrane

1989 ◽  
Vol 94 (2) ◽  
pp. 307-318
Author(s):  
S. Campbell ◽  
T.D. Allen ◽  
B.B. Moser ◽  
J.D. Aplin
Keyword(s):  
Electron Microscopy ◽  
Transmission Electron Microscopy ◽  
Cell Surface ◽  
Basal Lamina ◽  
Ammonium Hydroxide ◽  
Apical Surface ◽  
Lamina Densa ◽  
Transmission Electron ◽  
Extraction Pattern ◽  
Stromal Matrix

The organisation of extracellular matrix beneath the human amniotic epithelium was investigated in order that the co-ordinate synthesis of basal lamina and stroma by these cells could be better understood. Transmission electron microscopy of intact tissue confirmed that stromal matrix fibrils are located between the cell surface and the basal lamina, and also penetrate the lamina. The distribution of the supralaminal fibrils and their association with the lamina was further investigated by scanning electron microscopy (SEM) after removal of the overlying epithelium. Five complementary procedures were used to remove the cells from the underlying lamina. Trypsin-EDTA treatment caused the epithelial cells to retract or detach from the lamina. SDS or ammonium hydroxide was used to extract the epithelium, which was then removed by physical shearing. Transmission electron microscopy (TEM) confirmed that the lamina densa and supralaminal fibres were present after extraction by these agents. Incubation in CHAPS, a zwiterionic detergent, did not remove the epithelium but permitted exposure of the basal lamina by mechanical scoring. Extraction with boric acid followed by osmium tetroxide produced epithelial disruption and revealed the lamina and stroma in different areas. Although the extraction pattern was different in each case, all of the five methods confirmed that individual fibrils and fibril bundles are present on the apical surface of, and enter, the lamina densa. Examination of the stromal surface of the basal lamina after fracture revealed fibrils passing from the stroma into the lamina densa. We therefore suggest that, in this tissue, newly synthesised stromal matrix components appear in an assembled fibrillar form between the basal cell surface and the basal lamina before becoming associated with the sublaminal stroma.

scholarly journals Clostridium perfringens produces an adhesive pilus required for the pathogenesis of necrotic enteritis in poultry

2021 ◽  
Author(s):  
D. Lepp ◽  
Y. Zhou ◽  
S. Ojha ◽  
I. Mehdizadeh Gohari ◽  
J. Carere ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Transmission Electron Microscopy ◽  
Cell Surface ◽  
Clostridium Perfringens ◽  
Direct Evidence ◽  
Necrotic Enteritis ◽  
Null Mutant ◽  
Transmission Electron ◽  
Mutant Strains ◽  
Important Disease

Clostridium perfringens Type G strains cause necrotic enteritis (NE) in poultry, an economically important disease that is a major target of in-feed antibiotics. NE is a multifactorial disease, involving not only the critically-important NetB toxin, but also additional virulence and virulence-associated factors. We previously identified a C. perfringens chromosomal locus (VR-10B) associated with disease-causing strains that is predicted to encode a sortase-dependant pilus. In the current study, we sought to provide direct evidence for the production of a pilus by C. perfringens and establish its role in NE pathogenesis. Pilus structures in virulent C. perfringens strain CP1 were visualized by transmission electron microscopy (TEM) of immuno-gold labelled cells. Filamentous structures were observed extending from the cell surface in wild-type CP1, but not from isogenic pilin-null mutant strains. In addition, immuno-blotting of cell surface proteins demonstrated that CP1, but not the null mutant strains, produced a high molecular weight ladder-like pattern characteristic of a pilus polymer. Binding to collagen types I, II and IV was significantly reduced (Tukey’s; p<0.01) in all three pilin mutants compared to CP1, and could be specifically blocked by CnaA and FimA antisera, indicating that these pilins participate in adherence. Furthermore, both fimA and fimB null mutants were both severely attenuated in their ability to cause disease in an in vivo chicken NE challenge model. Together, these results provide the first direct evidence for the production of a sortase-dependant pilus by C. perfringens, and confirm its critical role in NE pathogenesis and collagen-binding. Importance In necrotic enteritis (NE), an intestinal disease of chickens, Clostridium perfringens cells adhere tightly to damaged intestinal tissue, but the factors involved are not known. We previously discovered a cluster of C. perfringens genes predicted to encode a pilus, a hair-like bacterial surface structure commonly involved in adherence. In the current study, we have directly imaged this pilus using transmission electron microscopy (TEM). We also show that inactivation of the pilus genes stops pilus production, significantly reducing the bacterium's ability to bind collagen and cause disease. Importantly, this is the first direct evidence for the production of a sortase-dependant pilus by C. perfringens, revealing a promising new target for developing therapeutics to combat this economically important disease.

scholarly journals Embedding cell monolayers to investigate nanoparticle-plasmalemma interactions at transmission electron microscopy

2019 ◽  
Vol 63 (1) ◽  
Author(s):  
Manuela Costanzo ◽  
Manuela Malatesta
Keyword(s):  
Electron Microscopy ◽  
Transmission Electron Microscopy ◽  
Cell Surface ◽  
Inexpensive Method ◽  
Biological Barrier ◽  
Cell Monolayers ◽  
Ultrastructural Evidence ◽  
Transmission Electron ◽  
Uptake Process ◽  
Intracellular Fate

Transmission electron microscopy is the technique of choice to visualize the spatial relationships between nanoconstructs and cells and especially to monitor the uptake process of nanomaterials. It is therefore crucial that the cell surface be preserved in its integrity, to obtain reliable ultrastructural evidence: the plasmalemma represents the biological barrier the nanomaterials have to cross, and the mode of membrane-nanoconstruct interaction is responsible for the intracellular fate of the nanomaterials. In this paper, we describe a simple and inexpensive method to process cell monolayers for ultrastructural morphology and immunocytochemistry, ensuring consistent preservation of the cell surface and of the occurring interactions with nanoparticles of different chemical composition.

Surface membrane regeneration in deciliated Tetrahymena

1983 ◽  
Vol 62 (1) ◽  
pp. 407-417
Author(s):  
N.E. Williams
Keyword(s):  
Electron Microscopy ◽  
Transmission Electron Microscopy ◽  
Membrane Proteins ◽  
Cell Surface ◽  
Surface Membrane ◽  
Membrane System ◽  
Ciliated Protozoa ◽  
Membrane Flow ◽  
Transmission Electron ◽  
Surface Marking

The induced synthesis of identified surface membrane proteins has been demonstrated in deciliated Tetrahymena. Cells in the process of regenerating cilia were also studied using transmission electron microscopy in order to obtain information on the deployment of new membrane at the cell surface. The results obtained suggest a pattern of membrane flow that includes the ‘pellicular alveoli’, a subsurface membrane system characteristically present in ciliated protozoa. The results of 125I surface-marking experiments were consistent with the notion that new membrane is added initially in non-ciliated regions, then subsequently flows laterally to cover regenerating cilia.

scholarly journals Novel protocol to observe the intestinal tuft cell using transmission electron microscopy

Biology Open ◽  
2022 ◽  
Author(s):  
Takuma Kozono ◽  
Miwa Tamura-Nakano ◽  
Yuki I. Kawamura ◽  
Takashi Tonozuka ◽  
Atsushi Nishikawa
Keyword(s):  
Electron Microscopy ◽  
Transmission Electron Microscopy ◽  
Specific Cell ◽  
Thin Sections ◽  
Cell Functions ◽  
Tuft Cell ◽  
Apical Side ◽  
Transmission Electron ◽  
Tuft Cells ◽  
Small Intestinal

Tuft cell is a chemosensory cell, a specific cell type sharing the taste transduction system with a taste cell on the tongue, of which the existence has been known in various tissues such as gastrointestinal tract, gall bladder, trachea, pancreatic duct, etc. To date, electron microscopic approaches have shown various morphological features of the tuft cell such as long and thick microvilli, tubulovesicular network at the apical side, prominent skeleton structures, etc. Recently, it has been reported that the small intestinal tuft cell functions to initiate type2 immunity in response to helminth infection. However, the mechanisms by which such distinguished structures are involved with the physiological functions are poorly understood. To address this question, the combination of physiological study regarding the tuft cells using genetic models and its morphological study using electron microscopy will be required. However, it is a challenge to observe tuft cells by electron microscopy due to their extremely low frequency on the epithelium. Therefore, in this paper, we suggest the advanced protocol to observe the small intestinal tuft cell efficiently by transmission electron microscopy using serial semi-thin sections on the Aclar film.

scholarly journals Immunomagnetic Purification of Colletotrichum lindemuthianum Appressoria

2000 ◽  
Vol 66 (8) ◽  
pp. 3464-3467 ◽  
Author(s):  
Katie A. Hutchison ◽  
Sarah E. Perfect ◽  
Richard J. O'Connell ◽  
Jonathan R. Green
Keyword(s):  
Electron Microscopy ◽  
Monoclonal Antibody ◽  
Transmission Electron Microscopy ◽  
Cell Surface ◽  
Cell Walls ◽  
Biochemical Analysis ◽  
Immunomagnetic Separation ◽  
Colletotrichum Lindemuthianum ◽  
Transmission Electron ◽  
Bean Anthracnose

ABSTRACT We developed a method to purify appressoria of the bean anthracnose fungus Colletotrichum lindemuthianum for biochemical analysis of the cell surface and to compare appressoria with other fungal structures. We used immunomagnetic separation after incubation of infected bean leaf homogenates with a monoclonal antibody that binds strongly to the appressoria. Preparations with a purity of >90% could be obtained. Examination of the purified appressoria by transmission electron microscopy showed that most had lost their cytoplasm. However, the plasma membrane was retained, suggesting that there is some form of attachment of this membrane to the cell wall. The purified appressoria can be used for studies of their cell surface, and we have shown that there are clear differences in the glycoprotein constituents of cell walls of appressoria compared with mycelium.

Studies of anionic sites on the cell surface of the amoeba Naegleria gruberi using cationized ferritin

1977 ◽  
Vol 28 (1) ◽  
pp. 133-149
Author(s):  
C.A. King ◽  
T.M. Preston
Keyword(s):  
Electron Microscopy ◽  
Transmission Electron Microscopy ◽  
Cell Surface ◽  
Binding Sites ◽  
Cationized Ferritin ◽  
Fluorescent Staining ◽  
Anionic Sites ◽  
Transmission Electron ◽  
Anionic Groups ◽  
Naegleria Gruberi

Interaction of cationized ferritin with the anionic groups on the cell surface of Naegleria was studied using transmission electron microscopy in conjunction with fluorescence microscopy. Most of the experiments involved the use of fluorescein-labelled cationized ferritin (FITC-CF) incubated with living amoebae. Initially the FITC-CF was located over the posterior two-thirds of the amoebae but the label was rapidly redistributed to form a cap at the posterior end in the region of the uroid; frequently this cap was shed. Pinosomes containing FITC-CF were clearly visible within the amoebae. Amoebae prefixed in glutaraldehyde were uniformly stained and did not show redistribution of the label. Exposure of live amoebae, previously incubated with cationized ferritin and allowed to cap, to fresh FITC-CF failed to produce fluorescent staining of the general cell surface, i.e. depletion of binding sites had occurred. The binding of the FITC-CF was not affected by pretreatment of the amoebae with neuraminidase or pronase. The possible nature of the anionic sites on the membrane is discussed.

Techniques for Transmission Electron Microscopy of Fiber-Matrix Interfaces in Aluminum Alloy-Boron Composites by Ion Erosio

1971 ◽  
Vol 29 ◽  
pp. 204-205
Author(s):  
G. G. Shaw
Keyword(s):  
Electron Microscopy ◽  
Transmission Electron Microscopy ◽  
Aluminum Alloy ◽  
Electron Beam ◽  
Pure Aluminum ◽  
Ion Milling ◽  
Transmission Electron ◽  
Fiber Matrix ◽  
Ion Erosion ◽  
Row Of Fibers

The morphology and composition of the fiber-matrix interface can best be studied by transmission electron microscopy and electron diffraction. For some composites satisfactory samples can be prepared by electropolishing. For others such as aluminum alloy-boron composites ion erosion is necessary.When one wishes to examine a specimen with the electron beam perpendicular to the fiber, preparation is as follows: A 1/8 in. disk is cut from the sample with a cylindrical tool by spark machining. Thin slices, 5 mils thick, containing one row of fibers, are then, spark-machined from the disk. After spark machining, the slice is carefully polished with diamond paste until the row of fibers is exposed on each side, as shown in Figure 1.In the case where examination is desired with the electron beam parallel to the fiber, preparation is as follows: Experimental composites are usually 50 mils or less in thickness so an auxiliary holder is necessary during ion milling and for easy transfer to the electron microscope. This holder is pure aluminum sheet, 3 mils thick.

Direct Observation of Heat Exchanger Deposits by Cross Sectioning

1967 ◽  
Vol 25 ◽  
pp. 364-365
Author(s):  
R. W. Anderson ◽  
D. L. Senecal
Keyword(s):  
Electron Microscopy ◽  
Transmission Electron Microscopy ◽  
Heat Exchanger ◽  
Direct Observation ◽  
Cross Sections ◽  
Preparation Method ◽  
Specimen Preparation ◽  
Flowing Water ◽  
Transmission Electron ◽  
Deposit Layer

A problem was presented to observe the packing densities of deposits of sub-micron corrosion product particles. The deposits were 5-100 mils thick and had formed on the inside surfaces of 3/8 inch diameter Zircaloy-2 heat exchanger tubes. The particles were iron oxides deposited from flowing water and consequently were only weakly bonded. Particular care was required during handling to preserve the original formations of the deposits. The specimen preparation method described below allowed direct observation of cross sections of the deposit layers by transmission electron microscopy.The specimens were short sections of the tubes (about 3 inches long) that were carefully cut from the systems. The insides of the tube sections were first coated with a thin layer of a fluid epoxy resin by dipping. This coating served to impregnate the deposit layer as well as to protect the layer if subsequent handling were required.

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