scholarly journals Adsorption of horseradish peroxidase, ovomucoid and anti-immunoglobulin to colloidal gold for the indirect detection of concanavalin A, wheat germ agglutinin and goat anti-human immunoglobulin G on cell surfaces at the electron microscopic level: a new method, theory and application.

1977 ◽  
Vol 25 (11) ◽  
pp. 1187-1200 ◽  
Author(s):  
W D Geoghegan ◽  
G A Ackerman
Keyword(s):  
Horseradish Peroxidase ◽  
Concanavalin A ◽  
Colloidal Gold ◽  
Wheat Germ Agglutinin ◽  
Wheat Germ ◽  
Indirect Detection ◽  
Electron Microscopic Level ◽  
Specific Cell ◽  
Major Protein ◽  
Electron Microscopic

A method is described for the adsorption of selected macromolecules to colloidal gold which is then used as an electron dense marker for the indirect detection of specific cell surface molecules. Membrane bound concanavalin A, which binds specific sugars on horseradish peroxidase, and wheat germ agglutinin, which binds specific sugars on ovomucoid are detected indirectly with gold labeled horseradish peroxidase and ovomucoid, respectively. Goat anti-human IgM on blood lymphocytes is detected with gold labeled rabbit anti-goat IgG. In the preparation of colloidal gold labeled proteins, the problems of flocculation of colloidal gold by proteins and nonadsorption of proteins to colloidal gold, are solved through a combination of concentration of protein and pH variable adsorption isotherms, which allows one to determine the conditions for adsorption of proteins to colloidal gold. Adsorption is pH dependent, the pH conditions correlating with the isoelectric point(s) of the major protein fraction(s); adsorption is influenced by interfacial tension, solubility and by the electrical charge on the molecules. Colloidal gold is inexpensive and preparation of a useful label is rapid, reproducible and the results easily quantitated from electron micrographs.

scholarly journals Influence of Concanavalin A, wheat germ agglutinin, and soybean agglutinin on the fusion of myoblasts in vitro.

1975 ◽  
Vol 67 (3) ◽  
pp. 826-834 ◽  
Author(s):  
H Den ◽  
D A Malinzak ◽  
H J Keating ◽  
A Rosenberg
Keyword(s):  
Concanavalin A ◽  
Binding Sites ◽  
Wheat Germ Agglutinin ◽  
Wheat Germ ◽  
Inhibitory Effect ◽  
Myoblast Fusion ◽  
Normal Serum ◽  
Specific Cell ◽  
Number Of Binding Sites

Although muscle cell fusion was shown to be an energy-requiring process, release of myoblasts from an EGTA fusion block could be accomplished with Earle's balanced salt solution (containing 1.8 mM Ca++) free of glucose or any other energy-produced metabolite. The effect of concanavalin A, abrin, and the lectins from wheat germ, soybean, and Lens culinaris on myoblast fusion was examined with synchronized myoblast cultures upon release from fusion block. At a concentration of 15 mug/ml, these lectins were found to inhibit the fusion process to the extent of 62%, 41%, 32%, 8%, and 19%, respectively. Concanavalin A inhibition could be prevented by alpha-methyl-D-mannoside. The inhibitory effect of all the lectins except abrin could be reversed by changing to the normal, serum-containing medium. The number of binding sites was 3.4 X 10(7), 6.1 X 10(7), and 1.7 X 10(6), respectively. Although myoblasts were found to have about twice as many binding sites for wheat germ agglutinin as for concanavalin A, concanavalin A was determined to be twice as effective as wheat germ agglutinin as an inhibitor of myoblast fusion. These findngs raise the possibility that specific cell surface glycoproteins may be an important factor in this process.

scholarly journals Lectin-binding sites as markers of Golgi subcompartments: proximal-to-distal maturation of oligosaccharides.

1983 ◽  
Vol 97 (4) ◽  
pp. 1243-1248 ◽  
Author(s):  
A M Tartakoff ◽  
P Vassalli
Keyword(s):  
Wheat Germ Agglutinin ◽  
Wheat Germ ◽  
Lectin Binding ◽  
Electron Microscopic Level ◽  
Asymmetric Distribution ◽  
Secretory Product ◽  
Electron Microscopic ◽  
Oligosaccharide Structure ◽  
Golgi Stack ◽  
Cellular Components

We investigated the subcellular sites of glycoprotein oligosaccharide maturation by using lectin conjugates to stain lightly-fixed, saponin-permeabilized myeloma cells. At the electron microscopic level, concanavalin A-peroxidase stains the cisternal space of the nuclear envelope, the rough endoplasmic reticulum, and cisternae along the proximal face of the Golgi stack. Conversely, wheat germ agglutinin-peroxidase stains cisternae along the distal face of the Golgi stack, associated vesicles, and the cell surface. These observations confirm the existence of two qualitatively distinct Golgi subcompartments, show that the lectin conjugates can be employed as relatively proximal or distal Golgi markers under conditions of excellent ultrastructural preservation, suggest that the asymmetric distribution of qualitatively distinct oligosaccharides is a property of underlying cellular components and not simply of the principal secretory product, and suggest that the oligosaccharide structure recognized by wheat germ agglutinin is attained during transport from the proximal toward the distal face of the Golgi stack.

scholarly journals Receptor-mediated gonadotropin action in the ovary. Inhibitory actions of concanavalin A and wheat-germ agglutinin on gonadotropin-stimulated cyclic AMP and progesterone responses in ovarian cells

1981 ◽  
Vol 200 (1) ◽  
pp. 153-159 ◽  
Author(s):  
S Azhar ◽  
K M Menon
Keyword(s):  
Cyclic Amp ◽  
Concanavalin A ◽  
Wheat Germ Agglutinin ◽  
Wheat Germ ◽  
Carbohydrate Chain ◽  
Inhibitory Effect ◽  
Dolichos Biflorus ◽  
Acetylneuraminic Acid ◽  
Ovarian Cells ◽  
Proline Incorporation

Pretreatment of ovarian cells with concanavalin A and wheat-germ agglutinin blocked the gonadotropin-induced cyclic AMP and progesterone responses and this effect was time- and concentration-dependent. Basal production of either cyclic AMP or progesterone, however, was not affected by treatment of cells with lectin. The effect of concanavalin A on gonadotropin-mediated cyclic AMP and progesterone responses was blocked by alpha-methyl D-mannoside and alpha-methyl d-glucoside. Similarly the inhibitory effect of wheat-germ agglutinin was reversed by N-acetyl-D-glucosamine. Pretreatment of ovarian cells with concanavalin A or wheat-germ agglutinin had no effect on protein synthesis in the ovary as monitored by [3H]proline incorporation studies. Concanavalin A and wheat-germ agglutinin did not affect steroid production in response to dibutyryl cyclic AMP and 8-bromo cyclic AMP, indicating that the inhibitory action of lectin was occurring at a step before cyclic AMP formation. Lectins specific for L-fucose, D-galactose and N-acetyl-D-galactosamine, gorse seed agglutinin, peanut agglutinin and Dolichos biflorus agglutinin respectively, did not interfere with gonadotropin-induced cyclic AMP and progesterone responses. The present studies suggest that gonadotropin receptors may be glycoprotein in nature or closely associated with glycoprotein structures with the carbohydrate chain containing N-acetyl-D-glucosamine, mannose and possibly N-acetylneuraminic acid.

Sign in / Sign up

Export Citation Format

Share Document