Immunophotoelectron microscopy of the cell surface and cytoskeleton.
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Animals↗
Biomedical subjects
K K Hedberg
Publications and source records attributed to K K Hedberg.
Silver-enhanced colloidal gold as a cell surface marker for photoelectron microscopy.
Colloidal gold labeling in conjunction with silver enhancement was investigated as a labeling technique for photoelectron microscopy (PEM). PEM uses UV-stimulated electron emission to image uncoated cell surfaces, and markers for cell surfaces need to be sufficiently photoemissive to be clearly visible against this background. Label contrast provided by 6 nm or 20 nm colloidal gold markers alone was compared to that provided by 6 nm markers after silver enhancement, using both direct and indirect labeling methods for fibronectin on human fibroblast cell surfaces. In all cases, details of the fibrillar fibronectin labeling distribution which were barely discernible before silver enhancement became highly visible against the cellular surface features. Two factors evidently contribute to the pronounced increase in label contrast with silver enhancement: (1) Increased particle size, which was documented by transmission electron microscopy, and (2) increased photoemission resulting from a silver coating on the enhanced gold markers, compared with the protein coating on the unenhanced gold markers. These data demonstrate that silver enhancement of colloidal gold labeling patterns in PEM images is a highly effective method for localization of specific sites on cell surfaces.
Cell Membrane↗
Early phorbol ester induced release of cell surface fibronectin: direct observation by photoelectron microscopy.
One of the early effects of the phorbol ester tumor promoter 12-0-tetradecanoylphorbol-13-acetate (TPA) on cultured normal fibroblasts is the release of fibronectin into the culture medium. Immunophotoelectron microscopy was used to follow the loss of fibronectin from the upper cell surface of normal human foreskin fibroblasts exposed to TPA. Fibronectin labeled with silver-enhanced colloidal gold-antibody conjugates appears in photoelectron images as streak- and network-like patterns of bright dots against the less photoemissive uncoated cell surface. Labeled fibronectin present beneath the culture is not detected due to the surface-specificity of this technique. Ten to 30 min of exposure to 100 ng/ml TPA in culture medium results in a readily visible decrease in upper cell surface fibronectin. In these experiments, 60 min of exposure to TPA releases nearly all upper cell surface fibronectin, leaving only occasional short streaks of label. In contrast, extracellular matrix fibronectin between cells is apparently more resistant to release and can still be seen in photoelectron images even when the upper cell surface appears to be fibronectin-free. This immunophotoelectron study shows the distribution of fibronectin on fibroblasts at high resolution and demonstrates that the initial fibronectin release resulting from TPA exposure is at the expense of preexisting cell-surface fibronectin. These results also illustrate the application of photoelectron microscopy as a useful technique in cell biology.
Cell Line↗