Mapping gold-labeled receptors on cell surfaces by backscattered electron imaging and digital image analysis: studies of the IgE receptor on mast cells.
The response of cells to signaling molecules such as hormones, growth factors, and immune mediators that bind to cell-surface receptors depends in part on the density and distribution of the relevant receptors. We have developed methods to map the distribution of IgE receptors on RBL-2H3 mast cells at high resolution in the scanning electron microscope (SEM). The key elements of our procedure are a new fixative that preserves receptor binding activity; a family of colloidal gold-conjugated probes that bind directly or indirectly to the IgE-receptor complex; an SEM with detectors for both secondary and backscattered electrons (to observe surface topography and gold particles, respectively); and an image processor that can average, digitize, and store these images. Topographical maps are generated by processing and superimposing the digitized images. The methods we describe can be applied to study the density and distribution of any membrane receptor that can be labeled with colloidal gold particles.
University of New Mexico School of Medicine, Albuquerque 87131.
R F
RF
Stump
J R
JR
Pfeiffer
M C
MC
Schneebeck
J C
JC
Seagrave
J M
JM
Oliver
D002462:Cell Membrane / Q000378:metabolism / Q000648:ultrastructure; D003432:Cross-Linking Reagents / Q000494:pharmacology; D004705:Endocytosis; D006046:Gold; D007089:Image Enhancement / Q000379:methods; D007091:Image Processing, Computer-Assisted; D007073:Immunoglobulin E / Q000378:metabolism*; D008024:Ligands; D008407:Mast Cells / Q000378:metabolism* / Q000502:physiology / Q000648:ultrastructure; D008855:Microscopy, Electron, Scanning / Q000379:methods*; D012834:Silver; D013205:Staphylococcal Protein A
D003432:Cross-Linking Reagents; D008024:Ligands; D013205:Staphylococcal Protein A; D007073:Immunoglobulin E; D012834:Silver; D006046:Gold