CONICET | Buscador de Institutos y Recursos Humanos

Quantum dots (QDs) posess unique properties (brightness, photostability, narrowband emission and broadband absorption), and excellent bio(chemical)compatibility for imaging structures and functions of living cells. QDs conjugated with ligands, are able to recognize and track multiple targets and visualize dynamic processes. Such QDs can be directed to precise cellular targets, detecting biomolecules with a sensitivity extending to the single molecule level. Thus, one can study the essential processes underlying the functions and regulation of living cells with probes providing "partial molecular derivatives", without which dissecting complex networks is impossible. Besides the brilliance and photostability of QDs that allow prolonged imaging and tracking of individul nanoparticles (NPs), other characteristics related to the local density and nature of molecular components can be exploited in cells and tissues for purposes other than mere detection. In particular, the systematic engineering of the molecular composition on the surface of QDs has been a central feature in numerous applications ranging beyond biosensing to cell delivery and release and to the activation of individual reactions and entire pathways.A widespread tendency is to consider the existence of multiple groups on the surface of NPs as disadvantageous in comparison with single attachment or reactive points, the rationale being that extensive conjugation may lead to a distribution of NP subpopulations differing in particle-label stoichiometries, to undesirable and unphysiological cross-linking, and other effects. Procedures for achieving precise control of the number of sites (down to single moieties) have been developed. We will maintain in this presentation, however, that under certain circumstances the presence of a large number of sites may offer distinct advantages. In fact, numerous studiesdemonstrate that multiple identical or different molecules on the surface of NPs can be exploited for for a wide variety of purposes, ranging from the control of optical or physical properties to the concurrent regulation of different functions. Commercially available NPs coated with a polymeric layer to isolate the core-shell structure from the aqueous environment have diameters of 15-20 nm and may extend to double these values if covered with PEG. Is this considerable size important such that QDs operate more as platforms rather than as single molecular entities? The distinction is subtle. For example, QDs have been shown to be useful as FRET donors and acceptors in numerous applications, although there have been relatively few reports of experiments based on imaging. In our own studies of cellular signalling mediated by growth factors and their receptors, QDs conjugated with ligands have revealed the existence of novel transport and trafficking mechanisms. In one instance, a biotinylated organic dye FRET acceptor served to demonstrate the existence of retrograde transport, but not endocytosis, of activated EGF receptor (EGFR), on filopodia, cellular extensions with a core of actin filaments. Figure X.1 depicts a strategy for using QDs attached to ligands as vehicles for specific targeting of cell surface receptors. In this case, QD-bound biotinylated EGF served to tag the EGF receptor and through this interaction, provided insight into the early stages of internalization of this prototypic family of receptor tyrosine kinases (RTKs).QD donors have been applied extensively in FRET-based assays of enzymatic activities, with the systematic introduction of multifunctionality being a major issue. The distance dependence of FRET operating via surface-bound or nearby small-molecule acceptors reflects the complex interplay between factors such as stoichiometry, spatial distribution and orientation, composition (passivation shell, capping moieties), and shape. (QDs emitting at > 600 nm tend to be non-spherical and demonstrate finite emission polarization).Optical properties such as fluorescence intensity or lifetime can also be tailored with FRET acceptors. The design of FRET based biosensors required for recording given states also requires control of the number of acceptors associated with each QD donor.