Single quantum dot tracking of membrane receptors.
The dynamics of membrane proteins in living cells has become a major issue to understand important biological questions such as chemotaxis, synaptic regulation, or signal transduction. The advent of semi-conductor quantum dots (QDs) has opened new perspectives for the study of membrane properties because these new nanomaterials enable measurements at the single molecule level with high signal-to-noise ratio. Probes used until now indeed encounter significant limitations: organic fluorophores and fluorescent proteins rapidly photobleach, whereas gold particles and latex beads, although more stable, are bulky and usually stain only one protein per experiment. In comparison, QDs are bright and photostable fluorescent probes with a size on the order of 10 nm can be used with standard immunochemical methods. We present the experimental protocols and methods of analysis which we used to investigate the dynamics of individual GABAA receptors in the axonal growth cone of spinal neurons in culture. Single QD tracking is nevertheless a general method, suitable to study many transmembrane proteins.
Bouzigues
Cédric
C
Laboratoire Kastler Brossel, Physics Department, Ecole Normale Supérieure, Paris, France.
Lévi
Sabine
S
Triller
Antoine
A
Dahan
Maxime
M
D000818:Animals; D001369:Axons / Q000737:chemistry / Q000378:metabolism; D002478:Cells, Cultured; D020439:Growth Cones / Q000737:chemistry / Q000378:metabolism; D007150:Immunohistochemistry; D008856:Microscopy, Fluorescence / Q000379:methods*; D009474:Neurons / Q000166:cytology / Q000378:metabolism; D045663:Quantum Dots; D051381:Rats; D011963:Receptors, GABA-A / Q000737:chemistry / Q000378:metabolism
D011963:Receptors, GABA-A