Gold labeling of cell monolayers grown on dextran beads.
Gold labeling of antigenic sites has become an increasingly useful tool in the study of cultured cell monolayers. If these monolayers are grown on flat substrates, major difficulties in both scanning (SEM) and transmission electron microscopy (TEM) specimen preparation and imaging may result. An alternate surface, that of dextran microcarrier beads, eliminates a majority of these difficulties and facilitates correlative TEM and SEM. The SEM procedure for using backscattered electron imaging requires the use of carbon planchets as the cell growth matrix to eliminate background signals. These planchets are expensive and are not an optimal cell-attachment matrix in that they result in loose and abnormally shaped cells. In contrast, the dextran beads were produced specifically for cell culture and, therefore, provide an excellent surface for growth. The beads have an average diameter of 100 microns, allowing attachment directly to aluminum stubs without signal generation from the aluminum to interfere with the gold signal. With TEM preparation, the monolayer poses the major disadvantage. Specimen preparation for thin sectioning is often preceded by extensive manipulation. In the microcarrier bead system, the beads are directly sectionable, and it is possible to cut five to eight full beads per thin section. This increase in cell surface makes quantification of gold labeling easier and also provides a more representative sampling of the monolayer. The ease of preparation, the decrease in reagents used (via cell pooling), and the ability to use one cell preparation for TEM and SEM make this procedure an ideal technique for gold labeling.
Hogan
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ME
Center for Electron Optics, Michigan State University, East Lansing 48824.
Hassouna
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HI
Klomparens
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KL
D000818:Animals; D003584:Cytological Techniques; D003911:Dextrans; D004730:Endothelium, Vascular / Q000166:cytology* / Q000648:ultrastructure; D006046:Gold; D008854:Microscopy, Electron; D008855:Microscopy, Electron, Scanning / Q000379:methods; D008863:Microspheres
D003911:Dextrans; D006046:Gold