Electron Microscopic Observations on the H Antigen Sites of Human Erythrocytes Using Ferritin Antibody Conjugates

We performed light and electron microscopic in situ hybridization, according to the same protocol and without pretreatment of sections, on Lowicryl- and LR Gold-embedded cells. Digoxigenin (DIG)- or biotin-labeled riboprobes were visualized by direct or indirect immunodetection using commercially available gold-antibody conjugates with 0.8-10-nm gold grains. At the ultrastructural level, the main findings were that DIG-labeled probes gave a slightly higher labeling intensity (grains per signal) than biotin. The direct detection method produced a more compact signal, which led to better resolution at medium and high magnifications. Labeling intensities of all gold grain sizes were essentially equal. Grain sizes of 5 nm and larger were highly preferable because available enhancement methods are unsatisfactory for ultrasmall grains. The optimized immunodetection protocols are suitable for double hybridization with two different probes and for combined hybridization and immunocytochemistry.

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Electron microscopic examination of the interaction between human erythrocytes and liposomes containing DC-Cholesterol

Bulletin of Experimental Biology and Medicine ◽

10.1007/bf02446035 ◽

1996 ◽

Vol 122 (1) ◽

pp. 724-727

Author(s):

Yu. S. Tarakhovskii

Keyword(s):

Microscopic Examination ◽

Electron Microscopic Examination ◽

Human Erythrocytes ◽

Electron Microscopic

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Electron Microscopic Detection of the Cryptantigen AHPhum (Friedenreich Antigen) on Human Erythrocytes by means of Gold-labelled Agglutinin from Helix Pomatia

Zeitschrift für Immunitätsforschung Immunobiology ◽

10.1016/s0340-904x(77)80060-2 ◽

1977 ◽

Vol 153 (5) ◽

pp. 450-456

Author(s):

M. Wagner ◽

B. Wagner

Keyword(s):

Helix Pomatia ◽

Human Erythrocytes ◽

Electron Microscopic ◽

Microscopic Detection

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THE PROPERTIES OF SPECIFIC STAINS FOR ELECTRON MICROSCOPY PREPARED BY THE CONJUGATION OF ANTIBODY MOLECULES WITH FERRITIN

The Journal of Cell Biology ◽

10.1083/jcb.9.3.519 ◽

1961 ◽

Vol 9 (3) ◽

pp. 519-537 ◽

Cited By ~ 198

Author(s):

S. J. Singer ◽

Anita F. Schick

Keyword(s):

Electron Scattering ◽

Specific Binding ◽

Electron Microscopic Examination ◽

Resolving Power ◽

Electron Microscopic ◽

Binding Properties ◽

Homologous Antigen ◽

Recent Developments ◽

Antibody Conjugates ◽

Antigen Antibody

In order to take full advantage of recent developments in the electron microscopic examination of cellular ultrastructure and composition, it is necessary to develop specific electron stains capable of identifying and localizing a wide variety of macromolecular components of cells. To this end, antibody conjugates have been prepared by chemically coupling the highly electron-scattering ferritin molecule to antibody. Antigen-antibody precipitations with these ferritin-antibody conjugates have demonstrated that under the appropriate conditions they retain the specific binding properties of the antibody from which they are prepared. An electron microscopic study has been made of aggregates of tobacco mosaic virus and its ferritin-conjugated antibody. The aggregates were prepared in solution and then sprayed onto specimen screens. The electron micrographs reveal that the conjugate specifically attached to, and delineated, the virus rods. The chemistry, structure, and resolving power of the ferritin-antibody conjugates, the specificity of their reactions with homologous antigen, and the nature of the problems to be faced in application of these conjugates to the study of the internal antigens of cells are discussed.

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The effect of EDTA, cations, and various buffers on the morphology of erythrocyte membranes: an electron-microscopic study

Blood ◽

10.1182/blood.v45.5.709.709 ◽

1975 ◽

Vol 45 (5) ◽

pp. 709-724 ◽

Cited By ~ 16

Author(s):

L Pinteric ◽

JF Manery ◽

IH Chaudry ◽

G Madapallimattam

Keyword(s):

Fine Structure ◽

Electron Microscope ◽

Microscopic Study ◽

Electron Microscopic Study ◽

Ethylenediaminetetraacetic Acid ◽

Human Erythrocytes ◽

Erythrocyte Membranes ◽

Electron Microscopic ◽

Osmotic Hemolysis ◽

Ionic Calcium

Abstract Membranes of human erythrocytes were prepared by stepwise osmotic hemolysis in Ca2+-free solutions. Examination with the electron microscope after negative staining showed some short, conelike protuberances on the surface of about 20 percent of the ghosts, while 80 percent were round, intact spheres. After Ca2+ treatment, all membranes were round and intact. After exposure to ethylenediaminetetraacetic acid (EDTA) (1.0 mM, pH 7.4), the entire ghost surface was covered with long, thin extrusions called stromalytic forms (about 460 per cell). Their sizes, shapes, and fine structure are described. Exposure to ionic calcium (1.4 times 10-minus 4M) abolished the EDTA-induced stromalytic forms. A second exposure to EDTA reversed this Ca2+ effect. ATP, like EDTA, produced stromalytic forms. EDTA- induced stromalytic forms were also abolished by Zn2+, La3+, and Nd3+ at concentrations of 1–5 times 10-minus 4 M. Mg2+ at 10-minus 2 M was ineffective. Ghosts were prepared by graded lysis in various buffers. Those prepared in phosphate were the most stable and provided consistent EDTA effects and Ca2+ reversal. Ghosts in Tris-HCl showed breakdown unless salt was added. Moderately satisfactory ghosts were also obtained in Hepes-NaOH buffer and salt.

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