Biologically compatible quantum dot (QD) nanoparticles are hybrid inorganic-organic materials with increasing
popularity as fluorescent probes for studying biological specimens. QDs have several advantageous optical features
compared to fluorescent dyes and they are electron-dense, allowing for correlated fluorescence and electron microscopic
imaging. Despite these features, widespread use of QDs as biological probes has generally been limited by the complex
chemistry required for their synthesis and the conjugation. In this work, we show that easily prepared quantum dot (QD)
probes provide excellent contrast for fluorescent confocal and environmental scanning electron microscopy (ESEM)
analysis of pure microbial cultures and microbial communities. Two conjugation strategies were employed in order to
specifically target the QDs to bacterial cell surfaces. The first was biotinylation of the bacteria followed by labeling with
commercially available QDs incorporating the high-affinity partner for biotin (QD-streptavidin). Second, we designed a
novel QD probe for Gram negative bacteria: QD-polymyxin B (PMB), which binds to lipopolysaccharide (LPS) in the
Gram negative cell wall. Pure cultures of Gram positive and Gram negative strains were used to illustrate that QDs
impart electron density and irradiation stability to the cells, and so no other preparation apart from QD labeling is
required. The techniques were then extended to a set of recently characterized microbial communities of perennial cold
springs in the Canadian High Arctic, which live in close association with unusual sulfur crystals. Using correlated
confocal and and ESEM, we were able to image these organisms in living samples and illustrate their relationship to the
In this work, we demonstrate the application of quantum dots (QDs) to several biologically relevant applications. QDs are synthesized by biological and organometallic routes and the relative merits of these methods are identified. Our results indicate that QDs can be functionalized and specifically targeted to both mammalian and bacterial cells. In the case of mammalian cells, they can be targeted to an engineered sodium channel for the purpose of sensing. In both mammalian and bacterial cells, the interaction with bioconjugated QDs can lead to phototoxicity due to the generation of reactive oxygen species (ROS).
Energy transfer between semiconductor nanoparticles (quantum dots) and energy donors or acceptors modulates fluorescence, thus serving as a visual indicator of the interaction. Careful choice of conjugate or capping groups can thus make these particles serve as sensors for specific biological processes and as tools for targeted cell killing. Challenges include creation of stable conjugates, delivery to specific cell populations and intracellular regions, and characterization of fluorescence modulation by energy transfer.
Semiconductor quantum dots (QDs) possess highly reactive electrons and holes after photoexcitation. The energy of these electrons and holes can be deliberately modulated by attaching the QD to an electron donor or acceptor. This eliminates (quenches) QD fluorescence, as well as affecting the ability of the QD to oxidize or reduce common biomolecules such as glutathione and DNA. This greatly alters the fluorescent properties and toxicity of such QDs
inside cells. In this work, we show that a specific electron donor, the neurotransmitter dopamine, yields redox-sensitive conjugates when attached to at least some colors of CdSe/ZnS QDs. The potential for the use of such conjugates as sensors, and the implications for enhanced toxicity in such conjugates are discussed.
The interaction between semiconductor nanocrystals (quantum dots) and biological structures and cells is strongly influenced by nanoparticle size, exposure to light, and surface cap or conjugate. Hydrophobic particles can insert into lipid bilayers, often resulting in membrane leakage. Oxidizing and reducing agents can photosensitize the quantum dot, yielding greater potential for cell damage; however, penetration into cells is seen only if the particle is specifically targeted to a receptor or antigen on the cell. When particles interact with DNA, oxidation can occur as measured by the presence of hydroxyguanine, preventing cellular replication. In this paper, several cell-free and whole-cell systems are presented to investigate the mechanisms for nanoparticle entry across lipid bilayers, the evolution of their surface composition with light and oxygen exposure, and their potential as targeted cytotoxic drugs and/or environmental hazards. Uptake into mammalian cells, Gram positive and Gram negative bacteria were compared and contrasted in order to identify important factors in nanoparticle uptake and toxicity. Simultaneously, Fourier transform infrared spectroscopy (FTIR) and time-correlated single photon counting (TCSPC) were used to track quantum dot surface degradation with time. Finally, a lipid bilayer system was used to investigate nanoparticle-membrane interactions. The advantages of this system are that its composition is fully known, so that the role of cell-surface receptors is eliminated, and recordings may be performed in the dark. These studies allowed for the formulation of preliminary models of quantum dot binding and entry that consider novel variables.