In vivo imaging using quantum-dot-conjugated probes.
This unit describes the use of quantum dots (QDs) for live-cell imaging and the use of QDs in flow cytometry for quantitative analysis of ligand binding constants and receptor density. Conventional fluorophores and visible fluorescent protein (VFP) constructs have allowed visualization of many cellular processes. However, organic and biomolecular fluorophores have limitations in their applications, due to their small Stokes' shift and tendency to photobleach during prolonged imaging. QDs have many advantages over conventional fluorophores, including high brightness and photostability, which make them an exceptional tool for live-cell imaging. There are a large variety of commercially available QDs with different surface reactivities and characteristics. The authors have limited the laboratory protocols presented here to the use of streptavidin-coupled QDs because this gives almost universal applicability to any cell surface receptor by coupling the ligand or antibody that recognizes the receptor to biotin and visualizing the complex by use of QDs.
S Lidke
Diane
D
University of New Mexico, Albuquerque, New Mexico, USA.
Nagy
Peter
P
J Arndt-Jovin
Donna
D
D000818:Animals; D000937:Antigen-Antibody Reactions; D001666:Binding Sites, Antibody; D019921:Biotinylation; D002478:Cells, Cultured / Q000648:ultrastructure; D005434:Flow Cytometry / Q000379:methods; D005456:Fluorescent Dyes / Q000032:analysis*; D006801:Humans; D007202:Indicators and Reagents; D008024:Ligands; D051379:Mice; D008856:Microscopy, Fluorescence / Q000295:instrumentation / Q000379:methods; D008863:Microspheres; D045663:Quantum Dots; D011956:Receptors, Cell Surface / Q000378:metabolism; D019809:Streptavidin
D005456:Fluorescent Dyes; D007202:Indicators and Reagents; D008024:Ligands; D011956:Receptors, Cell Surface; D019809:Streptavidin