Quantitative single particle tracking analysis of NGF retrograde transport

MARÍA M. ECHARTE, LUCIANA BRUNO, DONNA J. ARNDT-JOVIN,THOMAS M. JOVIN AND LÍA I. PIETRASANTA

FEBS LETTERS

Año: 2007 vol. 581 p. 2905 - 2913

0014-5793

NGF retrograde transport in living differentiated PC12 neurite-like processes was studied at thesingle molecule level using monobiotin-NGF (bNGF) coupled to streptavidin-quantum dots(QDs). These reagents retained the properties of unlabeled NGF with respect to the processes of differentiation, binding and internalization. Between 10 and 35% of the complexes movedtowards the cell body while the rest remained at or fluctuated about their initial position.Retrograde movement required the integrity of cytoskeletal microtubules. Quantitative singleparticle tracking analysis revealed a bidirectional step-like motion with a net component in theretrograde direction and a mean velocity of 0.054 ± 0.020 μm/s. Individual runs (i.e. a segment of uninterrupted displacement) had a mean velocity of ~0.15 μm/s at room temperature and anexponentially decaying distribution of run times. The photostability and small size of the QDspermit for the first time the extended real-time imaging of individual QD-bNGF-receptorcomplexes trafficking within neuronal-like cells.