scholarly journals mAb CZP-315.D9: An Antirecombinant Cruzipain Monoclonal Antibody That Specifically Labels the Reservosomes ofTrypanosoma cruziEpimastigotes

2014 ◽  
Vol 2014 ◽  
pp. 1-9 ◽  
Author(s):  
Cassiano Martin Batista ◽  
Lia Carolina Soares Medeiros ◽  
Iriane Eger ◽  
Maurilio José Soares
Keyword(s):  
Electron Microscopy ◽  
Monoclonal Antibody ◽  
Cysteine Proteinase ◽  
Igg Antibody ◽  
Western Blots ◽  
Myeloma Cells ◽  
Transmission Electron ◽  
Large Round ◽  
Endocytosis Pathway ◽  
Limiting Dilution

Reservosomes are large round vesicles located at the posterior end of epimastigote forms of the protozoanTrypanosoma cruzi, the etiological agent of Chagas disease. They are the specific end organelles of the endocytosis pathway ofT. cruzi, and they play key roles in nutrient uptake and cell differentiation. These lysosome-like organelles accumulate ingested macromolecules and contain large amounts of a major cysteine proteinase (cruzipain or GP57/51 protein). Aim of this study was to produce a monoclonal antibody (mAb) against a recombinantT. cruzicruzipain (TcCruzipain) that specifically labels the reservosomes. BALB/c mice were immunized with purified recombinant TcCruzipain to obtain the mAb. After fusion of isolated splenocytes with myeloma cells and screening, a mAb was obtained by limiting dilution and characterized by capture ELISA. We report here the production of a kappa-positive monoclonal IgG antibody (mAb CZP-315.D9) that recognizes recombinant TcCruzipain. This mAb binds preferentially to a protein with a molecular weight of about 50 kDa on western blots and specifically labels reservosomes by immunofluorescence and transmission electron microscopy. The monoclonal CZP-315.D9 constitutes a potentially powerful marker for use in studies on the function of reservosomes ofT. cruzi.

Acid-base effects on intestinal Na+ absorption and vesicular trafficking

2002 ◽  
Vol 283 (3) ◽  
pp. C971-C979 ◽  
Author(s):  
Alan N. Charney ◽  
Richard W. Egnor ◽  
Jesline Alexander-Chacko ◽  
Nicholas Cassai ◽  
Gurdip S. Sidhu
Keyword(s):  
Electron Microscopy ◽  
Apical Membrane ◽  
Ringer Solution ◽  
Vesicular Trafficking ◽  
Acid Base ◽  
Western Blots ◽  
Ph Values ◽  
Transmission Electron ◽  
Apical Membranes ◽  
Nhe Isoforms

We examined for vesicular trafficking of the Na+/H+ exchanger (NHE) in pH-stimulated ileal and CO2-stimulated colonic Na+absorption. Subapical vesicles in rat distal ileum were quantified by transmission electron microscopy at ×27,500 magnification. Internalization of ileal apical membranes labeled with FITC-phytohemagglutinin was assessed using confocal microscopy, and pH-stimulated ileal Na+ absorption was measured after exposure to wortmannin. Apical membrane protein biotinylation of ileal and colonic segments and Western blots of recovered proteins were performed. In ileal epithelial cells incubated in HCO[Formula: see text]/Ringer or HEPES/Ringer solution, the number of subapical vesicles, the relative quantity of apical membrane NHE isoforms 2 and 3 (NHE2 and NHE3, respectively), and apical membrane fluorescence under the confocal microscope were not affected by pH values between 7.1 and 7.6. Wortmannin did not inhibit pH-stimulated ileal Na+ absorption. In colonic epithelial apical membranes, NHE3 protein content was greater at a Pco 2 value of 70 than 21 mmHg, was internalized when Pco 2 was reduced, and was exocytosed when Pco 2 was increased. We conclude that vesicle trafficking plays no part in pH-stimulated ileal Na+absorption but is important in CO2-stimulated colonic Na+ absorption.

A MORPHOLOGICAL APPROACH TO THE MECHANISMS OF RISTOCETIN INDUCED PLATELET AGGLUTINATION(RIPA)

1987 ◽  
Author(s):  
G Escolar ◽  
J Monteagudo ◽  
N Villamor ◽  
M Garrido ◽  
R Castillo
Keyword(s):  
Electron Microscopy ◽  
Monoclonal Antibody ◽  
Platelet Rich Plasma ◽  
Human Platelets ◽  
Complex Interaction ◽  
Electron Dense Deposit ◽  
Transmission Electron ◽  
Dense Deposit ◽  
Morphological Approach ◽  
Adhesive Proteins

Changes in the morphology of human platelets induced by Ristocetin (RIPA) have been analyzed at ultraestructural level by means of a tannine acid procedure. Studies were completed with aggregation and binding experiments. Modificationsof these tests induced by apyrase,a monoclonal antibody (Mab) to GPIIb/IIIa and EDTA were also investigated.Transmission electron microscopy reveals that ristocetin precipitates adhesive proteins on plateletmembrane. An electron-dense deposit was noticeable within 20 secondsafter ristocetin was added. When experiments were carried out in theaggregometer cuvette under stirring, groups of platelets become activated, change shape, and finally aggregate releasing part of their content. The morphology of aggregates did not differ from those formed in the presence of ADP.Aggregation studies demonstrated that a Mab to GPIIb/IIIa modifies the extent and the rate of the aggregation curve when RIPA was performed in citrated platelet rich plasma (c-PRP). Apyrase modified the extent but not the slope of the curve. Neither the antibody nor apyrase modified RIPA when it was performed in PRP obtained in presence of EDTA. Binding experiments confirmed that I-vWF bound to platelets in presence of ristocetin was not modified by apyrase or anti-GPIIb/IIIa Mab. All these facts together suggest that RIPA,when performed in c-PRP besides reflecting the interaction of GPI with vWF, is also testing other mechanisms of the platelet function including exposure of GPIIb/IIIa complex, interaction of fibrinogen with this glycoprotein, and the contribution of the release reaction.

Delayed immunologic thrombocytopenia induced by abciximab

2004 ◽  
Vol 92 (10) ◽  
pp. 820-828 ◽  
Author(s):  
Gisèle Clofent-Sanchez ◽  
Catherine Jais ◽  
Emilse Bermejo ◽  
Lionel Leroux ◽  
Pierre Coste ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Monoclonal Antibody ◽  
Coronary Intervention ◽  
Inhibitory Effect ◽  
Immunogold Labeling ◽  
Antibody Immobilization ◽  
Igg Antibodies ◽  
Igg Antibody ◽  
Percutaneous Coronary ◽  
Thrombotic Complications

SummaryAbciximab is an anti-GPIIb-IIIa drug widely used to prevent thrombotic complications during percutaneous coronary intervention. We now report on the immunologic origin of thrombocytopenia developing between 7 and 12 days after the onset of abciximab infusion. Antibodies directed against abciximabcoated platelets were located in 5 patients with delayed thrombocytopenia, just as they were present in a patient whose platelet count fell within a few hours after receiving the drug. Abciximab-dependent IgG antibody was revealed in serum using control platelets in the monoclonal antibody immobilization of platelet antigens assay (MAIPA) performed with SZ22, a MoAb to GPIIb. The presence of IgG antibodies specific for platelets sensitized with abciximab was confirmed by flow cytometry. They were not located in 13 patients receiving abciximab but whose platelet counts remained stable. For three patients, antibodies were transient and their presence related to the extent of the thrombocytopenia. Surprisingly, antibodycontaining plasma from three patients induced abciximabdependent activation and aggregation of normal platelets, a finding confirmed by electron microscopy. Immunogold labeling revealed that abciximab was associated with platelets in the aggregate, suggesting that its inhibitory effect was overcome by the platelet stimulation. In summary, these results show that abciximab-dependent thrombocytopenia can be delayed and potentially prothrombotic.

Use of Transmission Electron Microscopy in Diagnosis of Acute Leukemias: A Prospective Study of Fifty Cases.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 4509-4509
Author(s):  
Tathagata Chatterjee ◽  
Manoranjan Mahapatra ◽  
Hara P. Pati ◽  
Inusha Panigrahi ◽  
Rajat Kumar ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Transmission Electron Microscopy ◽  
Bone Marrow Aspiration ◽  
Acute Leukemias ◽  
Ultrastructural Cytochemistry ◽  
Ultrastructural Studies ◽  
Transmission Electron ◽  
Large Round ◽  
Hypercellular Marrow ◽  
A Prospective Study

Abstract Ultrastructural studies have contributed substantially to the understanding of the cellular morphology of the acute leukemias. We present here 50 cases (aged 3-55 years, M: F-32: 18) of acute leukemias, which were studied for morphology, conventional cyto chemistry, immunophenotyping, and transmission electron microscopy (TEM) including ultrastructural cytochemistry using myeloperoxidase (MPO) and platelet peroxidase activity. TEM morphology using ultrastructural cytochemistry of MPO helped diagnose three cases of acute myeloid leukemia with minimal myeloid differentiation (AML M0). The blasts showed large round nuclei, 1–2 nucleoli, chromatin with peripheral condensation and abundant mitochondria. Of total of 5 cases of acute promyelocytic leukemias (APML), all had strong Sudan Black, MPO, dual esterase positivity; one case was non-specific esterase positive and sensitive to fluoride. On TEM, this unusual case was identified to be microgranular variant of APML. TEM morphology and ultrastructural cytochemistry using platelet peroxidase helped diagnose 3 cases of AML M7 (acute megakaryocytic leukemia), 2 cases of acute biphenotypic leukemias and also in differentiating one case of acute proerythroblastic leukemia (AML M6b) from 3 cases of AML M6a or acute erythroleukemia. Thus, TEM is helpful in differentiating further the subgroups of AML-M5 and AML-M6, in identifying the microgranular variant of APML, and in confirming the diagnosis of AML-M0 and biphenotypic leukemia. Also, in cases with very hypercellular marrow and with associated myelofibrosis, where the bone marrow aspiration gives low cell count, TEM and ultrastructural cytochemistry are a valuable aid to arriving at a accurate diagnosis. Characteristics of acute leukemia cases (n=50) FAB Subtype Morphology, Conventional Cytochemistry and Immunophenotyping Transmission Electron Microscopy (?) = doubt in diagnosis AML-M0 3 (?) 3 AML-M1and M2 11 11 AML-M3 4, 1(?) 5: Hypergranular-4, hypogranular-1 AML-M4 5 5 AML-M5 5 5a-4, 5b-1 AML-M6 4 6a-3 (erythroleukemia), 6b-1 AML-M7 3 3 Biphenotypic 2 (?) 2 ALL-L1 8 8 ALL-L2 4 4

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.

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.

Phase Transformations in Ti-7w/oMo-3w/oMe Alloy

1974 ◽  
Vol 32 ◽  
pp. 514-515
Author(s):  
S. Fujishiro
Keyword(s):  
Electron Microscopy ◽  
Mechanical Properties ◽  
Transmission Electron Microscopy ◽  
Tensile Strength ◽  
Mechanical Processing ◽  
Ray Method ◽  
X Ray ◽  
Transmission Electron ◽  
Water Quench ◽  
Alpha Beta

The mechanical properties of three titanium alloys (Ti-7Mo-3Al, Ti-7Mo- 3Cu and Ti-7Mo-3Ta) were evaluated as function of: 1) Solutionizing in the beta field and aging, 2) Thermal Mechanical Processing in the beta field and aging, 3) Solutionizing in the alpha + beta field and aging. The samples were isothermally aged in the temperature range 300° to 700*C for 4 to 24 hours, followed by a water quench. Transmission electron microscopy and X-ray method were used to identify the phase formed. All three alloys solutionized at 1050°C (beta field) transformed to martensitic alpha (alpha prime) upon being water quenched. Despite this heavily strained alpha prime, which is characterized by microtwins the tensile strength of the as-quenched alloys is relatively low and the elongation is as high as 30%.

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