Measuring the Molecular Scale Dynamics of Protein Receptor Endocytic Trafficking in NeurAL Cells Using Quantum Dot BioCONJugate

Protein trafficking is critical in neurons since neurons must orchestrate the movement of a plethora of discrete intracellular signaling proteins from the cell body to the ends of their axons, over distances that may span up to several meters. A problem in studying protein trafficking has been a lack of tools to visualize the movement of discrete proteins inside live neurons, in real time. An integrated understanding of endocytic trafficking at the level of single or small numbers of receptor complexes inside live cells is currently hampered by technical limitations. Here, we develop and apply quantum dot QD bioconjugates for imaging discrete receptor endocytic events inside live neural cells. QD probes can bind with specific cognate receptors consequently cell signaling cascades to regulate neural sprouting. Furthermore, QD-receptor complexes are internalized by cells dynamically traffic discrete receptor bound QDs on their membrane surface as well as along vast distances along intracellular microtubule tracks of neural processes. Using single particle tracking and immuno-colocalization, we illustrate and validate the use of QD-receptor complexes for imaging receptor trafficking at synchronized time points after QD-receptor binding and internalization. The unique value of these probes is illustrated by new dynamic observations: 1) that endocytosis proceeds at strikingly regulated fashion, and 2) that diffusive and active forms of transport inside cells are rapid and efficient. QDs are powerful intracellular probes that can provide investigators with new capabilities and fresh insight for studying endocytic receptor signaling events, in real time, and at the resolution of single or small numbers of receptors in live cell