Abstract
Markers for immuno-scanning electron microscopy had been, so far, selected for easy identification based on their distinctive shape (for example, haemocyanin, bacteriophage T4). These markers were always viewed in the secondary electron imaging (SEI) mode. Their size was not posing much of a problem of resolution for commercially available SEM, but was likely to prevent good labeling efficiency, due to steric hindrance phenomena. Higher labeling efficiency necessitates the use of markers of smaller size which, unfortunately, can rarely be unambiguously recognized in topographical SE images.To correlate antigenic distribution with small cell surface structures like microvilli or coated pits, markers of smaller size (i.e., in the 20 nm range) must be used and clearly identified on the surface of well preserved cells. With the availability of colloidal gold particles complexed with various ligands this became possible, particularly if the labeled cells are viewed in the backscattered imaging (BEI) mode of the SEM, therefore basing the identification of the marker on its atomic number contrast rather than on its topographical contrast.
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