Nanogold as a specific marker for electron cryotomography.
While electron cryotomography (ECT) provides "molecular" resolution, three-dimensional images of unique biological specimens, sample crowdedness, and/or resolution limitations can make it difficult to identify specific macromolecular components. Here we used a 1.4 nm Nanogold cluster specifically attached to the Fc fragment of IgG to monitor its interaction with the neonatal Fc receptor (FcRn), a membrane-bound receptor that transports IgG across cells in acidic intracellular vesicles. ECT was used to image complexes formed by Nanogold-labeled Fc bound to FcRn attached to the outer surface of synthetic liposomes. In the resulting three-dimensional reconstructions, 1.4 nm Nanogold particles were distributed predominantly along the interfaces where 2:1 FcRn-Fc complexes bridged adjacent lipid bilayers. These results demonstrate that the 1.4 nm Nanogold cluster is visible in tomograms of typically thick samples (approximately 250 nm) recorded with defocuses appropriate for large macromolecules and is thus an effective marker.
He
Yongning
Y
Division of Biology, California Institute of Technology, 114-96, 1200 East California Blvd., Pasadena, CA 91125, USA.
Jensen
Grant J
GJ
Bjorkman
Pamela J
PJ
D000818:Animals; D015415:Biomarkers / Q000737:chemistry; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D055032:Electron Microscope Tomography / Q000379:methods*; D006046:Gold / Q000737:chemistry*; D015395:Histocompatibility Antigens Class I / Q000737:chemistry; D007141:Immunoglobulin Fc Fragments / Q000737:chemistry; D007074:Immunoglobulin G / Q000737:chemistry; D008081:Liposomes / Q000737:chemistry; D053768:Metal Nanoparticles / Q000737:chemistry / Q000648:ultrastructure*; D011485:Protein Binding; D011961:Receptors, Fc / Q000737:chemistry; D013194:Staining and Labeling
D015415:Biomarkers; D015395:Histocompatibility Antigens Class I; D007141:Immunoglobulin Fc Fragments; D007074:Immunoglobulin G; D008081:Liposomes; D011961:Receptors, Fc; D006046:Gold; C100688:Fc receptor, neonatal