scholarly journals A novel approach for scanning electron microscopy of colloidal gold-labeled cell surfaces.

1984 ◽  
Vol 99 (1) ◽  
pp. 53-57 ◽  
Author(s):  
E de Harven ◽  
R Leung ◽  
H Christensen

A method is described for the use of scanning electron microscopy on the surface of gold-labeled cells. It includes the use of 45- or 20-nm colloidal gold marker conjugated with Staphylococcal protein A. The marker is best recognized on the basis of its atomic number contrast by using the backscattered electron imaging mode of the scanning electron microscope. When the backscattered electron signal is mixed with the secondary electron signal, an optimum correlation between the distribution of the labeled sites and the cell surface structures is demonstrated. The method is illustrated by its application to the identification of human circulating granulocytes. Its good resolution, high contrast, and good labeling efficiency offers a promising approach to the specific localization of cell surface antigenic sites labeled with particles of colloidal gold.

1984 ◽  
Vol 52 (02) ◽  
pp. 102-104 ◽  
Author(s):  
L J Nicholson ◽  
J M F Clarke ◽  
R M Pittilo ◽  
S J Machin ◽  
N Woolf

SummaryA technique for harvesting mesothelial cells is described. This entails collagenase digestion of omentum after which the cells can be cultured. The technique has been developed using the rat, but has also been successfully applied to human tissue. Cultured rat mesothelial cells obtained in this way have been examined by scanning electron microscopy. Rat mesothelial cells grown on plastic film have been exposed to blood in an in vitro system using a Baumgartner chamber and have been demonstrated to support blood flow. No adhering platelets were observed on the mesothelial cell surface. Fibroblasts similarily exposed to blood as a control were washed off the plastic.


1985 ◽  
Vol 40 (7-8) ◽  
pp. 539-550 ◽  
Author(s):  
Margarete Borg

Abstract The labeling of immunocomplexes for scanning electron microscopy (SEM) is a fairly new technique, and the various procedures, that have been proposed, have not yet been compared. Such comparative evaluation was performed with Candida protease as a target antigen. This secretory enzyme of the opportunistic yeast Candida albicans can be localized on the surface of fungal blastopores and mycelia, both after growth in proteinaceous medium and upon infection of murine peritoneal macrophages. The presence of the protease antigen was confirmed by immunofluorescence and by immunoperoxidase-light microscopy. The decoration of protease - anti protease complexes for SEM was attempted with colloids derived from the immunoperoxidase reaction, by the immunogold technique, and by antibodies linked to beads of synthetic polymers (polystyrene, polymethacrylate, polyacrolein). In addition, inactivated Staphylococcus aureus was used, which binds to antibodies through its protein-A. The high resolution by SEM of surface structures was matched only by the colloid based decoration techniques. All conjugates with beads suffered from inconsistent binding, which did not correspond with the distribution of the surface antigen. The comparatively best result with beads was obtained with polystyrene (Latex). Colloid based techniques in addition allow for critical point drying, which cannot be applied to synthetic beads in the usual manner.


1976 ◽  
Vol 71 (1) ◽  
pp. 314-322 ◽  
Author(s):  
R Molday ◽  
R Jaffe ◽  
D McMahon

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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