A scanning electron microscope study of lectin receptors on neuroblastoma cells

The organization and redistribution of concanavalin A (Con A), Ricinus communis agglutinin I (RCA I) and wheat germ agglutinin (WGA) receptors on mouse neuroblastoma cells was analyzed using lectins conjugated to fluorescent polymeric microspheres as visual markers for scanning electron microscopy (SEM) and fluorescent light microscopys. Results indicate that all three lectins are internalized by these cells but the patterns of redistribution for Con A differ from those of RCA I and WGA.Labeling of prefixed undifferentiated and differentiated neuroblastoma cells indicated that Con A, WGA and RCA I receptors were densely and uniformly distributed over the cell surface and neurites. Cells treated with lectin for 5-10 minutes at 37°C, washed free of excess reagent and maintained for up to 15 minutes in buffer exhibited uniform labeling patterns.

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Concanavalin a and wheat germ agglutinin receptors on dictyostelium: Their visualization by scanning electron microscopy with microspheres

The Journal of Cell Biology ◽

10.1083/jcb.71.1.314 ◽

1976 ◽

Vol 71 (1) ◽

pp. 314-322 ◽

Cited By ~ 28

Author(s):

R Molday ◽

R Jaffe ◽

D McMahon

Keyword(s):

Electron Microscopy ◽

Scanning Electron Microscopy ◽

Cell Surface ◽

Wheat Germ ◽

Cellular Interactions ◽

Con A ◽

Stages Of Development ◽

Lectin Receptors ◽

Plant Lectins ◽

Scanning Electron

The cellular slime mold, Dictyostelium discoideum, is a convenient model for studying cellular interactions during development. Evidence that specific cell surface components are involved in cellular interactions during its development has been obtained by Gerisch and co-workers (1, 2) using immunological techniques. Smart and Hynes (3) have shown that a cell surface protein can be iodinated on cells in aggregation phase, but not in vegetative phase, by the lactoperoxidase procedure. Recently, McMahon et al. (4), and Hoffman and McMahon have demonstrated, by SDS gel electrophoresis, considerable differences in cell surface proteins and glycoproteins of plasma membranes isolated from cells at different stages of development. Plant lectins have also been used to monitor changes in cell surface properties of D. discoideum cells during development. Weeks and co-workers (5, 6) have detected differences in the binding and agglutination of cells by concanavalin A (Con A). Gillette and Filosa (7) have shown that Con A inhibits cell aggregation and prematurely induces cyclic AMP phosphodiesterase. Capping of Con A receptors has also been reported (8). Reitherman et al. (9) have recently reported that agglutination of cells by several plant lectins and the slime mold agglutination, discoidin, changes during development. Such studies indicate that differences in surface properties exist for cells at various stages of development. However, owing to the uncertainties in the factors which contribute to lectin-induced cell agglutination (10), the molecular basis for these observations remain to be determined. In this study, we have used microspheres (11-14) coupled to either Con A or wheat germ agglutinin (WGA) as visual markers to study by scanning electron microscopy the topographical distribution of lectin receptors on D. discoideum cells fixed at different stages of development. We also describe the effect of labeling on the distribution of lectin receptors and on the morphology of the cell surface.

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Lectin-binding sites on the plasma membranes of rabbit spermatozoa: Changes in surface receptors during epididymal maturation and after ejaculation

The Journal of Cell Biology ◽

10.1083/jcb.74.3.950 ◽

1977 ◽

Vol 74 (3) ◽

pp. 950-962 ◽

Cited By ~ 125

Author(s):

GL Nicolson ◽

N Usui ◽

R Yanagimachi ◽

H Yanagimachi ◽

Smith JR

Keyword(s):

Binding Sites ◽

Plasma Membranes ◽

Ricinus Communis ◽

Lectin Binding ◽

Similar Degree ◽

Con A ◽

Lectin Receptors ◽

Surface Receptors ◽

Epididymal Spermatozoa ◽

Lectin Binding Sites

Modifications in rabbit sperm plasma membranes during epididymal passage and after ejaculation were investigated by used of three lectins: concanavalin A (Con A); Ricinus communis I (RCA(I)); and wheat germ agglutinin (WGA). During sperm passage from caput to cauda epididymis, agglutination by WGA drastically decreased, and agglutination by RCA(I) slightly decreased, although agglutination by Con A remained approximately unchanged. After ejaculation, spermatozoa were agglutinated to a similar degree or slightly less by Con A, WGA, and RCA(I), compared to cauda epididymal spermatozoa. Ultrastructural examination of sperm lectin-binding sites with ferritin- lectin conjugates revealed differences in the densities of lectin receptors in various sperm regions, and changes in the same regions during epididymal passage and after ejaculation. Ferritin-RCA(I) showed abrupt changes in lectin site densities between acrosomal and postacrosomal regions of sperm heads. The relative amounts of ferritin-RCA(I) bound to heads of caput epididymal or ejaculated spermatozoa. Tail regions were labeled by ferritin RCA(I) almost equally on caput and cauda epididymal spermatozoa, but the middle-piece region of ejaculated spermatozoa was slightly more densely labeled than the principal-piece region, and these two regions on ejaculated spermatozoa were labeled less than on caput and cuada epididymal spermatozoa. Ferritin-WGA densely labeled the acrosomal region of caput epididymal spermatozoa, although labeling of cauda epidiymal spermatozoa was relatively sparse except in the apical area of the acrosomal region. Ejaculated spermatozoa bound only a few molecules of ferritin-WGA, even at the highest conjugate concentrations used. Caput epididymal, but not cauda epididymal or ejaculated spermatozoa, bound ferritin-WGA in the tail regions. Dramatic differences in labeling densities during epididymal passage and after ejaculation were not found with ferritin-Con A.

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Ultrastructural localization of lectin receptors on the luminal and abluminal aspects of brain micro-blood vessels.

Journal of Histochemistry & Cytochemistry ◽

10.1177/34.2.3753715 ◽

1986 ◽

Vol 34 (2) ◽

pp. 251-261 ◽

Cited By ~ 59

Author(s):

A W Vorbrodt ◽

D H Dobrogowska ◽

A S Lossinsky ◽

H M Wisniewski

Keyword(s):

Blood Vessels ◽

Ricinus Communis ◽

Helix Pomatia ◽

Ultrastructural Localization ◽

Con A ◽

Gold Complexes ◽

Entire Cross Section ◽

Lectin Receptors ◽

Helix Pomatia Agglutinin ◽

Lowicryl K4m

Lectin- or glycoprotein-colloidal gold complexes were used for detection of specific monosaccharide residues in mouse brain micro-blood vessels (MBVs). The lectins tested recognize the following residues: beta-D-galactosyl (Ricinus communis agglutinin-120, RCA-1), alpha-N-acetylgalactosaminyl (Helix pomatia agglutinin, HPA), alpha-D-mannosyl and alpha-D-glucosyl (Concanavalin A, Con A), sialoglycoconjugates (Limax flavus agglutinin, LFA), N-acetylglucosaminyl and sialyl (wheat germ agglutinin, WGA), and alpha-L-fucosyl (Ulex europeus agglutinin, UEA-1). Use of these lectin-gold complexes and ultrathin sections of Lowicryl K4M-embedded tissue makes it possible to gain insights into localization of lectin receptors in the entire cross-section of MBV walls. Receptors for all lectins, except UEA-1, were found on both luminal and abluminal fronts of the endothelial cells (ECs). Differential labeling of luminal and abluminal fronts of ECs with some lectins (Con A, HPL) is considered to reflect the polarity of the endothelium. Some differences noted in the distribution of lectin receptors in the wall of representatives of three types of MBVs (capillaries, arterioles, and venules) are thought to be associated with different functions performed by the above-mentioned segments of the microvasculature in maintenance of the blood-brain barrier.

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Effect on glutaraldehyde fixation on lectin-mediated agglutination of mouse leukaemia cells

Journal of Cell Science ◽

10.1242/jcs.21.3.579 ◽

1976 ◽

Vol 21 (3) ◽

pp. 579-594

Author(s):

W.J. Van Blitterswijk ◽

E.F. Walborg ◽

C.A. Feltkamp ◽

H.A. Hilkmann ◽

P. Emmelot

Keyword(s):

Ricinus Communis ◽

Single Cells ◽

Cell Aggregation ◽

Cell Counting ◽

Cell Aggregates ◽

Glutaraldehyde Fixation ◽

Shear Forces ◽

Con A ◽

Lectin Receptors ◽

Free Surfaces

The effect of glutaraldehyde fixation on lectin-mediated agglutination of murine leukaemia (GRSL) cells was investigated using 2 assay methods which differed in the shear forces to which the agglutinated cells were subjected. First, lectin and cells were allowed to interact under conditions in which shear forces were minimized and the degree of agglutination was evaluated microscopically by the appearance and size of the cell aggregates. This assay demonstrated that concanavalin A (con A)-, wheat germ agglutinin (WGA)- or Ricinus communis agglutinin I (RCAI)-mediated cytoagglutination was unaffected (WGA and RCAI) or somewhat enhanced (con A) by prior fixation of the cells with glutaraldehyde. Secondly, an electronic particle counter was used to measure the disappearance of single cells and concomitant appearance of cell aggregates as a function of the lectin concentration. In this assay, in which the aggregated cells are subjected to significant shear forces during dilution and cell counting, agglutination of GRSL cells by each of the 3 lectins was drastically inhibited by prior fixation of the cells with glutaraldehyde. This assay also demonstrated enhanced nonlectin-induced cell aggregation after fixation. In both cytoagglutination assays about the same lectin concentration was required for threshold agglutination of unfixed cells. Comparatively, the results of the 2 cytoagglutination assays indicate that a fraction of the lectin-mediated bonds between unfixed cells is shear resistant and that fixation of the cells either weakens these bonds or inhibits their formation. Morphologically, cells prefixed with glutaraldehyde were sperical at all lectin concentrations, with a continuous dense distribution of cell surface-bound con A, labelled directly with haemocyanin or indirectly using the peroxidase-diaminobenzidine reaction. Unfixed cells showed angular and toadstool-shaped deformations, especially at the highest lectin concentrations, the agglutinating surfaces being flattened against each other over extended areas. The distribution of con A label was continuous and dense between the apposed surfaces and discontinuous on free surfaces. In the presence of con A the free surfaces of prefixed cells exhibited more microvilli than the surfaces of non-prefixed cells. These results favour the view that fixation prevents the formation of shear-resistant, lectin-mediated bonds between cells, not by restricting the lateral mobility of lectin receptors, but by impairing the apposition of rigid cell surfaces.

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Distribution and mobility of lectin receptors on synaptic membranes of identified neurons in the central nervous system.

The Journal of Cell Biology ◽

10.1083/jcb.71.2.487 ◽

1976 ◽

Vol 71 (2) ◽

pp. 487-496 ◽

Cited By ~ 80

Author(s):

P Kelly ◽

C W Cotman ◽

C Gentry ◽

G L Nicolson

Keyword(s):

Granule Cells ◽

Ricinus Communis ◽

Postsynaptic Membrane ◽

Synaptic Membranes ◽

Con A ◽

Lectin Receptors ◽

Mature Neuron ◽

The Central Nervous System ◽

Restricted Mobility ◽

Topographic Distribution

The distribution and mobility of concanavalin A (Con A) and Ricinus communis agglutinin (RCA) receptors (binding sites) on the external surfaces of Purkinje, hippocampal pyramidal, and granule cells and their attached boutons were studied using ferritin-lectin conjugates. Dendritic fields of these cells were isolated by microdissection and gently homogenized. Cell fragments and pre- and postsynaptic membranes were labeled with the ferritin-lectin conjugates at a variety of temperatures, and the distribution of lectin receptors was determined by electron microscopy. Both classes of these lectin receptors were concentrated at nearly all open and partially open postsynaptic junctional membranes of asymmetric-type synapses on all three neuron types. Con A receptors were most concentrated at the junctional membrane region, indicating that the mature neuron has a specialized nonrandom organization of carbohydrates on its outer surface. Lectin receptors located on postsynaptic junctional membranes appeared to be restricted in their mobility compared to similar classes of receptors on extrajunctional membrane regions. Labeling with ferritin-RCA and -Con A at 37 degrees C produced clustering of lectin receptors on nonjunctional surfaces; however, Con A and RCA receptors retained their nonrandom topographic distribution on the postsynaptic junctional surface. The restricted mobility of lectin receptors was an inherent property of the postsynaptic membrane since the presynaptic membrane was absent. It is proposed that structures in the postsynaptic density may be transmembrane-linked to postsynaptic receptors and thereby determine topographic distribution and limit diffusion of specialized synaptic molecules. Speicalized receptor displays may play an important role in the formation and maintenance of specific synaptic contacts.

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A Scanning Electron Microscope Study of Isolated Guard Cells

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s042482010007271x ◽

1973 ◽

Vol 31 ◽

pp. 454-455 ◽

Cited By ~ 1

Author(s):

P. Dayanandan ◽

P. B. Kaufman

Keyword(s):

Electron Microscope ◽

Electron Microscope Study ◽

Three Dimensional ◽

Guard Cells ◽

Scanning Electron Microscope Study ◽

Psilotum Nudum ◽

Subsidiary Cells ◽

Welwitschia Mirabilis ◽

Cycas Revoluta ◽

Scanning Electron

A three dimensional appreciation of the guard cell morphology coupled with ultrastjuctural studies should lead to a better understanding of their still obscure dynamics of movement. We have found the SEM of great value not only in studies of the surface details of stomata but also in resolving the structures and relationships that exist between the guard and subsidiary cells. We now report the isolation and SEM studies of guard cells from nine genera of plants.Guard cells were isolated from the following plants: Psilotum nudum, four species of Equisetum, Cycas revoluta, Ceratozamia sp., Pinus sylvestris, Ephedra cochuma, Welwitschia mirabilis, Euphorbia tirucalli and Allium cepa.

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