Hedi Mattoussi

Long-term multiple color imaging of live cells using quantum dot bioconjugates

Luminescent quantum dots (QDs)—semiconductor nanocrystals—are a promising alternative to organic dyes for fluorescence-based applications. We have developed procedures for using QDs to label live cells and have demonstrated their use for long-term multicolor imaging of live cells. The two approaches presented are (i) endocytic uptake of QDs and (ii) selective labeling of cell surface proteins with QDs conjugated to antibodies. Live cells labeled using these approaches were used for long-term multicolor imaging. The cells remained stably labeled for over a week as they grew and developed. These approaches should permit the simultaneous study of multiple cells over long periods of time as they proceed through growth and development.

Electrical, optical, and structural properties of indium-tin-oxide thin films for organic light-emitting devices

High-quality indium–tin–oxide (ITO) thin films (200–850 nm) have been grown by pulsed laser deposition (PLD) on glass substrates without a postdeposition annealing treatment. The structural, electrical, and optical properties of these films have been investigated as a function of target composition, substrate deposition temperature, background gas pressure, and film thickness. Films were deposited from various target compositions ranging from 0 to 15 wt % of SnO2 content. The optimum target composition for high conductivity was 5 wt % SnO2+95 wt % In2O3. Films were deposited at substrate temperatures ranging from room temperature to 300 °C in O2 partial pressures ranging from 1 to 100 mTorr. Films were deposited using a KrF excimer laser (248 nm, 30 ns full width at half maximum) at a fluence of 2 J/cm2. For a 150-nm-thick ITO film grown at room temperature in an oxygen pressure of 10 mTorr, the resistivity was 4×10−4 Ω cm and the average transmission in the visible range (400–700 nm) was 85%. For a 170-n...

Tracking metastatic tumor cell extravasation with quantum dot nanocrystals and fluorescence emission-scanning microscopy

Metastasis is an impediment to the development of effective cancer therapies. Our understanding of metastasis is limited by our inability to follow this process in vivo. Fluorescence microscopy offers the potential to follow cells at high resolution in living animals. Semiconductor nanocrystals, quantum dots (QDs), offer considerable advantages over organic fluorophores for this purpose. We used QDs and emission spectrum scanning multiphoton microscopy to develop a means to study extravasation in vivo. Although QD labeling shows no deleterious effects on cultured cells, concern over their potential toxicity in vivo has caused resistance toward their application to such studies. To test if effects of QD labeling emerge in vivo, tumor cells labeled with QDs were intravenously injected into mice and followed as they extravasated into lung tissue. The behavior of QD-labeled tumor cells in vivo was indistinguishable from that of unlabeled cells. QDs and spectral imaging allowed the simultaneous identification of five different populations of cells using multiphoton laser excitation. Besides establishing the safety of QDs for in vivo studies, our approach permits the study of multicellular interactions in vivo.

Electroluminescence from heterostructures of poly(phenylene vinylene) and inorganic CdSe nanocrystals

Electroluminescence (EL) and photoluminescence (PL) from heterostructure thin films made of organic poly (phenylene vinylene), PPV, and inorganic semiconductor CdSe nanocrystals are investigated. In these devices, the organic PPV structure is built next to an indium tin oxide anode, using the technique of molecular layer-by-layer sequential adsorption, and serves primarily as the hole transport layer. The inorganic layer, adjacent to an Al electrode, is made of spin cast CdSe nanocrystals, passivated with either organic groups or with a wider band gap semiconductor, e.g., ZnS in the present case. We find that the electroluminescence signal is almost exclusively generated within the inorganic layer, with a very weak contribution from the PPV layer at higher applied voltage. The performance of these heterostructure devices is influenced by the thickness of the dot layer. Lifetime tests reveal promising stability, with devices operating continuously over 50–100 h. Values of the external quantum efficiency, η...

Quantum-dot/dopamine bioconjugates function as redox coupled assemblies for in vitro and intracellular pH sensing

The use of semiconductor quantum dots (QDs) for bioimaging and sensing has progressively matured over the past decade. QDs are highly sensitive to charge-transfer processes, which can alter their optical properties. Here, we demonstrate that QD-dopamine-peptide bioconjugates can function as charge-transfer coupled pH sensors. Dopamine is normally characterized by two intrinsic redox properties: a Nernstian dependence of formal potential on pH and oxidation of hydroquinone to quinone by O(2) at basic pH. We show that the latter quinone can function as an electron acceptor quenching QD photoluminescence in a manner that depends directly on pH. We characterize the pH-dependent QD quenching using both electrochemistry and spectroscopy. QD-dopamine conjugates were also used as pH sensors that measured changes in cytoplasmic pH as cells underwent drug-induced alkalosis. A detailed mechanism describing the QD quenching processes that is consistent with dopamines inherent redox chemistry is presented.

Indium tin oxide thin films for organic light-emitting devices

High-quality indium tin oxide (ITO) thin films (150–200 nm) were grown on glass substrates by pulsed laser deposition (PLD) without postdeposition annealing. The electrical, optical, and structural properties of these films were investigated as a function of substrate temperature, oxygen pressure, and film thickness. PLD provides very uniform ITO films with high transparency (⩾85% in 400–700 nm spectrum) and low electrical resistivity (2–4×10−4 Ω cm). The Hall mobility and carrier density for a 170-nm-thick film deposited at 300 °C are 29 cm2/V s and 1.45×1021 cm−3, respectively. Atomic force microscopy measurements of the ITO films indicated that their root-mean-square surface roughness (∼5 A) is superior to that (∼40 A) of commercially available ITO films deposited by sputtering. ITO films grown at room temperature by PLD were used to study the electroluminescence (EL) performance of organic light-emitting devices. The EL performance was comparable to that measured with commercial ITO anodes.

Delivering quantum dots into cells: strategies, progress and remaining issues

The use of semiconductor quantum dots (QDs) in biological sensing and labeling continues to grow with each year. Current and projected applications include use as fluorescent labels for cellular labeling, intracellular sensors, deep-tissue and tumor imaging agents, sensitizers for photodynamic therapy, and more recently interest has been sparked in using them as vectors for studying nanoparticle-mediated drug delivery. Many of these applications will ultimately require the QDs to undergo targeted intracellular delivery, not only to specific cells, but also to a variety of subcellular compartments and organelles. It is apparent that this issue will be critical in determining the efficacy of using QDs, and indeed a variety of other nanoparticles, for these types of applications. In this review, we provide an overview of the current methods for delivering QDs into cells. Methods that are covered include facilitated techniques such as those that utilize specific peptide sequences or polymer delivery reagents and active methods such as electroporation and microinjection. We critically examine the benefits and liabilities of each strategy and illustrate them with selected examples from the literature. Several important related issues such as QD size and surface coating, methods for QD biofunctionalization, cellular physiology and toxicity are also discussed. Finally, we conclude by providing a perspective of how this field can be expected to develop in the future.

Luminescent quantum dots as platforms for probing in vitro and in vivo biological processes

In this report we review some of the recent progress made for enhancing the biocompatibility of luminescent quantum dots (QDs) and for developing targeted bio-inspired applications centered on live cell imaging and sensing. We start with a detailed analysis of the surface functionalization strategies developed thus far, and discuss their effectiveness for providing long term stability of the quantum dots in biological media, to changes in pH and to added electrolytes. We then discuss the available conjugation techniques to couple QDs to a variety of biological receptors and compare their effectiveness. In particular, we highlight the implementation of new strategies such as the use of copper-free cyclo-addition reaction (CLICK) chemistry and chemo-selective ligation. We then discuss the advances made for intracellular delivery where ideas such as receptor-driven endocytosis and uptake promoted by cell penetrating peptides are used. We then describe a few representative examples where QDs have been used to investigate specific cell biology processes. Such processes include binding of QDs conjugated to the nerve growth factor to membrane specific receptors and intracellular uptake, tracking of membrane protein at the single molecule level, and recognition of ligand bound QDs by T cell receptors. We conclude by discussing issues of toxicity associated with the use of QDs in biology.

Use of quantum dots for live cell imaging

. Despite their considerable advantages in live cell imaging, organic fluorophores are subject to certain limitations. Fluorescent quantum dots (QDs) are inorganic fluorescent nanocrystals that overcome many of these limitations and provide a useful alternative for studies that require long-term and multicolor imaging of cellular and molecular interactions

Sensing Caspase 3 Activity with Quantum Dot-Fluorescent Protein Assemblies

We demonstrate the use of a hybrid fluorescent protein semiconductor quantum dot (QD) sensor capable of specifically monitoring caspase 3 proteolytic activity. mCherry monomeric red fluorescent protein engineered to express an N-terminal caspase 3 cleavage site was ratiometrically self-assembled to the surface of QDs using metal-affinity coordination. The proximity of the fluorescent protein to the QD allows it to function as an efficient fluorescence resonance energy transfer acceptor. Addition of caspase 3 enzyme to the QD-mCherry conjugates specifically cleaved the engineered mCherry linker sequence, altering the energy transfer with the QD and allowing quantitative monitoring of proteolytic activity. Inherent advantages of this sensing approach include bacterial expression of the protease substrate in a fluorescently appended form, facile self-assembly to QDs, and the ability to recombinantly modify the substrate to target other proteases of interest.