scholarly journals The 17β-oestradiol dehydrogenase of pig endometrial cells is localized in specialized vesicles

1993 ◽  
Vol 290 (3) ◽  
pp. 777-782 ◽  
Author(s):  
J Adamski ◽  
B Husen ◽  
F Marks ◽  
P W Jungblut
Keyword(s):  
Electron Microscopy ◽  
Monoclonal Antibodies ◽  
Immunogold Electron Microscopy ◽  
The Novel ◽  
Electron Dense Material ◽  
Endometrial Cells ◽  
Functional Aspects ◽  
Cytoplasmic Vesicles ◽  
Similar Appearance ◽  
High Degree

Two monoclonal antibodies against the 17 beta-oestradiol dehydrogenase of pig endometrial cells have been used in localization studies with immunogold electron microscopy. The antibodies attach both to a fraction of dehydrogenase-rich cytoplasmic vesicles isolated from homogenates and to vesicles of similar appearance in cells. The vesicles are filled with electron-dense material. Their tagging intensity indicates a high degree of specialization. Endometrial cells from mature animals contain a host of dehydrogenase vesicles, and cells from prepubertal animals only a few. Functional aspects of the novel organelle are discussed.

Definition of individual components within the cytoskeleton of Trypanosoma brucei by a library of monoclonal antibodies

1989 ◽  
Vol 93 (3) ◽  
pp. 491-500 ◽  
Author(s):  
A. Woods ◽  
T. Sherwin ◽  
R. Sasse ◽  
T.H. MacRae ◽  
A.J. Baines ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Monoclonal Antibodies ◽  
Trypanosoma Brucei ◽  
Immunofluorescence Microscopy ◽  
Complex Mixtures ◽  
Immunogold Electron Microscopy ◽  
Basal Bodies ◽  
Tubulin Isotypes ◽  
Definition Of ◽  
Attachment Zone

The detergent-insoluble T. brucei cytoskeleton consists of several morphologically distinct regions and organelles, many of which are detectable only by electron microscopy. We have produced a set of monoclonal antibodies that define each structural component of this highly ordered cytoskeleton. The monoclonal antibodies were selected by cloning of hybridomas produced from mice injected with complex mixtures of proteins of either the cytoskeleton itself or salt extracts thereof. Four antibodies define particular tubulin isotypes and locate the microtubules of the axoneme and sub-pellicular array; two antibodies recognize the flagellum attachment zone; one recognizes the paraflagellar rod and another the basal bodies. Finally, one antibody defines a detergent-insoluble component of the nucleus. The antigens detected by each monoclonal antibody have been analysed by immunofluorescence microscopy, immunogold electron microscopy and Western blotting.

scholarly journals Rab4 Regulates Formation of Synaptic-like Microvesicles from Early Endosomes in PC12 Cells

2001 ◽  
Vol 12 (11) ◽  
pp. 3703-3715 ◽  
Author(s):  
Heidi de Wit ◽  
Yael Lichtenstein ◽  
Regis B. Kelly ◽  
Hans J. Geuze ◽  
Judith Klumperman ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Pc12 Cells ◽  
Early Endosome ◽  
Small Gtpase ◽  
Immunogold Electron Microscopy ◽  
Cell Periphery ◽  
Early Endosomes ◽  
Cytoplasmic Vesicles ◽  
Important Site

Early endosomes in PC12 cells are an important site for the formation of synaptic-like microvesicles and constitutive recycling vesicles. By immunogold electron microscopy, the small GTPase rab4 was localized to early endosomes and numerous small vesicles in the cell periphery and Golgi area of PC12 cells. Overexpression of GTPase-deficient Q67Lrab4 increased the number of early endosome-associated and cytoplasmic vesicles, whereas expression of GDP-bound S22Nrab4 significantly increased the length of early endosomal tubules. In parallel, Q67Lrab4 induced a shift in rab4, VAMP2, and TfR label from early endosomes to peripheral vesicles, whereas S22Nrab4 increased early endosome labeling of all three proteins. These observations were corroborated by early endosome budding assays. Together, our data document a thus far unrecognized role for rab4 in the formation of synaptic-like microvesicles and add to our understanding of the formation of constitutive recycling vesicles from early endosomes.

scholarly journals Monoclonal Antibodies to a Subfraction of Merino Wool High-tyrosine Proteins

1986 ◽  
Vol 39 (4) ◽  
pp. 341 ◽  
Author(s):  
DR Hewish ◽  
PW French
Keyword(s):  
Electron Microscopy ◽  
Monoclonal Antibodies ◽  
Immunosorbent Assay ◽  
Immunofluorescent Staining ◽  
Immunogold Electron Microscopy ◽  
Enzyme Linked Immunosorbent Assay ◽  
Specific Location ◽  
Merino Wool

Monoclonal antibodies were prepared which react with members of the high-tyrosine type proteins from Merino wool. Specificity was confirmed by the use of Western transfer immunoassays and by enzyme-linked immunosorbent assay on purified fractions. Immunofluorescent staining of sections of wool follicles using the antibodies showed that the proteins were present in the developing wool shaft but that staining was asymmetric, indicating specific location of the proteins in the orthocortex of the fibres. Immunogold-electron microscopy confirmed that one of the antibodies bound to the keratin microfibril bundles.

scholarly journals Orientation within the Exosporium and Structural Stability of the Collagen-Like Glycoprotein BclA of Bacillus anthracis

2005 ◽  
Vol 187 (15) ◽  
pp. 5310-5317 ◽  
Author(s):  
Jeremy A. Boydston ◽  
Ping Chen ◽  
Christopher T. Steichen ◽  
Charles L. Turnbough
Keyword(s):  
Electron Microscopy ◽  
Escherichia Coli ◽  
Monoclonal Antibodies ◽  
Bacillus Anthracis ◽  
Triple Helix ◽  
Thermal Denaturation ◽  
Sodium Dodecyl ◽  
Basal Layer ◽  
Structural Data ◽  
Immunogold Electron Microscopy

ABSTRACT Bacillus anthracis spores, which cause anthrax, are enclosed by an exosporium consisting of a basal layer and an external hair-like nap. The filaments of the nap are composed of BclA, a glycoprotein containing distinct N-terminal (NTD) and C-terminal (CTD) domains separated by an extended collagen-like central region. In this study, we used immunogold electron microscopy to show that the CTD of BclA forms the distal end of each filament of the hair-like nap, indicating that the NTD is attached to the basal layer. Ten randomly chosen anti-BclA monoclonal antibodies, raised against spores or exosporium, reacted with the CTD, consistent with its exterior location. We showed that recombinant BclA (rBclA), encoded by the B. anthracis Sterne strain and synthesized in Escherichia coli, forms a collagen-like triple helix as judged by collagenase sensitivity and circular dichroism spectroscopy. In contrast, native BclA in spores was resistant to collagenase digestion. Thermal denaturation studies showed that the collagen-like region of rBclA exhibited a melting temperature (T m ) of 37°C, like mammalian collagen. However, rBclA trimers exhibited T m values of 84°C and 95°C in buffer with and without sodium dodecyl sulfate, respectively. CTD trimers exhibited the same T m values, indicating that the high temperature and detergent resistances of rBclA were due to strong CTD interactions. We observed that CTD trimers are resistant to many proteases and readily form large crystalline sheets. Structural data indicate that the CTD is composed of multiple beta strands. Taken together, our results suggest that BclA and particularly its CTD form a rugged shield around the spore.

scholarly journals Use of immunogold electron microscopy and monoclonal antibodies in the identification of nuclear substructures.

1984 ◽  
Vol 32 (7) ◽  
pp. 757-765 ◽  
Author(s):  
C V Clevenger ◽  
A L Epstein
Keyword(s):  
Electron Microscopy ◽  
Monoclonal Antibodies ◽  
Ultrastructural Localization ◽  
Immunoblot Analysis ◽  
Immunogold Electron Microscopy ◽  
Supramolecular Organization ◽  
Nuclear Antigens ◽  
Interchromatin Granules ◽  
Cycle Dependent ◽  
Cytochemical Technique

A cytochemical technique for the ultrastructural localization of unique nuclear antigens is reported. Using a post-embedding indirect immunogold labeling procedure, nuclear antigens in electron-dense regions of the nucleus are localized with a minimum of nonspecific staining. Using this technique and indirect immunofluorescence, a panel of antinuclear monoclonal antibodies is shown to recognize preferentially cell cycle-dependent nuclear substructures. The antigenic domains recognized include specific regions in condensed chromatin, interchromatin granules, euchromatin, and chromosomes. The specificity of antigen recognition is demonstrated with qualitative and quantitative immunogold electron microscopy and immunoblot analysis. These results reveal the existence of previously undefined supramolecular organization within the nucleus and demonstrate the utility of the immunogold procedure when monoclonal antibodies are used.

Tissue section lifting for electron microscopy

1978 ◽  
Vol 36 (2) ◽  
pp. 86-87
Author(s):  
Adrian F. van Dellen
Keyword(s):  
Infectious Disease ◽  
Electron Microscopy ◽  
Tissue Culture ◽  
Organ System ◽  
Surrounding Tissue ◽  
Paraffin Sections ◽  
Surface Areas ◽  
Specific Determination ◽  
High Degree ◽  
Accuracy And Repeatability

The morphologic pathologist may require information on the ultrastructure of a non-specific lesion seen under the light microscope before he can make a specific determination. Such lesions, when caused by infectious disease agents, may be sparsely distributed in any organ system. Tissue culture systems, too, may only have widely dispersed foci suitable for ultrastructural study. In these situations, when only a few, small foci in large tissue areas are useful for electron microscopy, it is advantageous to employ a methodology which rapidly selects a single tissue focus that is expected to yield beneficial ultrastructural data from amongst the surrounding tissue. This is in essence what "LIFTING" accomplishes. We have developed LIFTING to a high degree of accuracy and repeatability utilizing the Microlift (Fig 1), and have successfully applied it to tissue culture monolayers, histologic paraffin sections, and tissue blocks with large surface areas that had been initially fixed for either light or electron microscopy.

Electron Microscopy in Molecular Biology

1967 ◽  
Vol 25 ◽  
pp. 2-3
Author(s):  
Cecil E. Hall
Keyword(s):  
Electron Microscopy ◽  
Molecular Biology ◽  
Noise Level ◽  
Molecular Structures ◽  
Material Structure ◽  
The Arts ◽  
Shadow Casting ◽  
Electron Image ◽  
High Degree ◽  
Very High

The visualization of organic macromolecules such as proteins, nucleic acids, viruses and virus components has reached its high degree of effectiveness owing to refinements and reliability of instruments and to the invention of methods for enhancing the structure of these materials within the electron image. The latter techniques have been most important because what can be seen depends upon the molecular and atomic character of the object as modified which is rarely evident in the pristine material. Structure may thus be displayed by the arts of positive and negative staining, shadow casting, replication and other techniques. Enhancement of contrast, which delineates bounds of isolated macromolecules has been effected progressively over the years as illustrated in Figs. 1, 2, 3 and 4 by these methods. We now look to the future wondering what other visions are waiting to be seen. The instrument designers will need to exact from the arts of fabrication the performance that theory has prescribed as well as methods for phase and interference contrast with explorations of the potentialities of very high and very low voltages. Chemistry must play an increasingly important part in future progress by providing specific stain molecules of high visibility, substrates of vanishing “noise” level and means for preservation of molecular structures that usually exist in a solvated condition.

scholarly journals Rapid and simultaneous purification of aflatoxin B1, zearalenone and deoxynivalenol using their monoclonal antibodies and magnetic nanoparticles

2021 ◽  
Author(s):  
Hyuk-Mi Lee ◽  
Hwan-Goo Kang
Keyword(s):  
Monoclonal Antibodies ◽  
Magnetic Nanoparticles ◽  
Aflatoxin B1 ◽  
Animal Feed ◽  
Immunoaffinity Chromatography ◽  
The Novel ◽  
Buffer Solution ◽  
Purification Method ◽  
Recovery Rates ◽  
Single Spike

AbstractTo develop a new simple and simultaneous purification method for mycotoxins in feeds and grains, magnetic nanoparticles (MNPs) conjugated with monoclonal antibodies (mAbs) against mycotoxins were used to separate aflatoxin B1 (AFB1), zearalenone (ZEA) and deoxynivalenol (DON). For a single spike of each mycotoxin into the buffer solution (16% MeOH in PBS), mean recoveries were 93.1–95.0% for AFB1 (5–20 ng/mL spiked), 87.2–96.0% for ZEA (125–500 ng/mL spiked) and 75.2–96.9% for DON (250–1,000 ng/mL spiked) by HPLC and ELISA. Recoveries of AFB1 (20 ng/mL) and ZEA (500 ng/mL) simultaneously spiked into the buffer solution were 87.0 and 99.8%, respectively. Recovery rates of AFB1/DON and DON/ZEA spiked simultaneously were 86.2%/76.6% and 92.0%/86.7%, respectively, at concentrations of 20 ng/mL AFB1, 500 ng/mL ZEA, and 1,000 ng/mL DON. Recoveries using the novel mAb–MNP conjugated system in a buffer solution simultaneously spiked with AFB1, ZEA and DON were 82.5, 94.6 and 73.4%, respectively. Recoveries of DON in animal feed were 107.7–132.5% at concentrations of 250–1,000 ng/g spiked in feed. The immunoaffinity chromatography (IAC) clean-up method was compared with the purification method using novel mAb–MNP. After fortification of animal feed with AFB1 (5, 10 and 20 ng/g feed) and ZEA (125, 250 and 500 ng/g feed), AFB1 and ZEA were purified using both the methods. In the case of the novel mAb-MNP conjugated system, mean recoveries for AFB1 were 89.4, 73.1 and 88.3% at concentrations of 5, 10 and 20 ng/g feed, respectively. For ZEA, mean recoveries were 86.7, 85.9 and 79.1% at concentrations of 125, 250 and 500 ng/g, respectively. For IAC purification, recoveries were 42.9–45.1% for AFB1 and 96.8–103.2% for ZEA. In conclusion, the present purification method using monoclonal antibodies conjugated to MNPs can be used for simple and simultaneous purification of mycotoxins from feed and maize.

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