Biomedical Engineering Reference
In-Depth Information
postsynaptic receptors? Traditional radiolabelling assays provide good sensitivity
and quantitative information on ligand-receptor binding; however radioactive
probes lack spatial and temporal information. Fluorescent or nanoparticle tagged
ligands would provide spatial information; however, at present the availability of
such probes is limited. The variety of QD bioconjugation methods offer good potential
for designing neuroactive ligand-conjugated QD probes to study receptor-mediated
activity and track drug binding and uptake in pharmaceutical and therapeutic
questions.
Initial studies using ligand-conjugated probes have shown that these probes bind
to their specifi c receptors without exhibiting signifi cant steric hindrance and pro-
vide high spatial resolution. Lidke et al. conjugated QDs to epidermal growth factor
(EGF) and monitored its binding to erbB1 receptors in CHO cells (Lidke et al.
2004
). Appending these receptors with enhanced GFP (eGFP) enabled simulta-
neous tracking of the ligand-receptor complex and its endocytosis into the cell.
Dynamic cellular processes, such as membrane ruffl ing, were observed prior to the
uptake of EGF-QD/erbB1-eGFP complexes and retrograde transport of these recep-
tors was observed in the fi lopodia of A431 cells.
The binding of nerve growth factor (NGF) to QDs to activate TrkA receptors in
PC-12 cells was studied by Vu et al. (
2005
). Short-term exposure to beta-NGF con-
jugated QDs, revealed specifi c QD binding to TrkA receptors, followed by internal-
ization of the ligand-receptor complexes. Long-term studies (3-5 days) demonstrated
that the NGF-QDs after binding to TrkA receptors, remained bioactive and were
capable of mimicking NGF activity as exhibited by extensive neurite sprouting.
Rosenthal et al. used QDs to gain more insight into transporter localization
and regulation (Rosenthal et al.
2002
; Tomlinson et al.
2005
) . They showed that
serotonin-conjugated QDs were not effective in inducing a current from the serotonin-3
receptors but were able to block the serotonin-transporter by binding to a site which
either overlaps with an antagonist site or is the serotonin-binding site itself. The
serotonin-QDs enabled direct visualization of the serotonin transporter protein on
the surface of transfected HEK cells and staining was completely eliminated on pre-
incubation with an antagonist.
These early studies demonstrate that QDs can be used as sensitive probes to
visualize ligand/drug traffi cking and study the signaling pathways of ligands of
interest. These results are encouraging as they show that QDs can be developed as
new tools to track effi cacy of ligand-receptor binding and the fate of ligand-receptor
complexes.
8.3
Concurrent Quantum Dots Fluorescence and Electron
Microscopy Imaging of Neural Ultrastructures
An advantage of QD probes is that they can be used as concurrent fl uorescent and
electron dense electron microscopy (EM) probes, thus giving access to additional
information in the same biological preparation. Dahan et al. performed single-receptor
