Quantum Dot−Fluorescent Protein Pairs as Novel Fluorescence Resonance Energy Transfer Probes

Fluorescence resonance energy transfer (FRET) characteristics, including the efficiency, donor−acceptor distance, and binding strength of six fluorescent protein (FP)−quantum dot (QD) pairs were quantified, demonstrating that FPs are efficient acceptors for QD donors with up to 90% quenching of QD fluorescence and that polyhistidine coordination to QD core−shell surface is a straightforward and effective means of conjugating proteins to commercially available QDs. This provides a novel approach to developing QD-based FRET probes for biomedical applications

Tiny Grains Give Huge Gains: Nanocrystal-Based Signal Amplification for Biomolecule Detection

Nanocrystals, despite their tiny sizes, contain thousands to millions of atoms. Here we show that the large number of atoms packed in each metallic nanocrystal can provide a huge gain in signal amplification for biomolecule detection. We have devised a highly sensitive, linear amplification scheme by integrating the dissolution of bound nanocrystals and metal-induced stoichiometric chromogenesis, and demonstrated that signal amplification is fully defined by the size and atom density of nanocrystals, which can be optimized through well-controlled nanocrystal synthesis. Further, the rich library of chromogenic reactions allows implementation of this scheme in various assay formats, as demonstrated by the iron oxide nanoparticle linked immunosorbent assay (ILISA) and blotting assay developed in this study. Our results indicate that, owing to the inherent simplicity, high sensitivity and repeatability, the nanocrystal based amplification scheme can significantly improve biomolecule quantification in both laboratory research and clinical diagnostics. This novel method adds a new dimension to current nanoparticle-based bioassays

Coating Optimization of Superparamagnetic Iron Oxide Nanoparticles for High T₂ Relaxivity

We describe a new method for coating superparamagnetic iron oxide nanoparticles (SPIOs) and demonstrate that, by fine-tuning the core size and PEG coating of SPIOs, the T2 relaxivity per particle can be increased by >200-fold. With 14 nm core and PEG1000 coating, SPIOs can have T2 relaxivity of 385 s−1 mM−1, which is among the highest per-Fe atom relaxivities. In vivo tumor imaging results demonstrated the potential of the SPIOs for clinical applications

Self-Assembly of Phospholipid–PEG Coating on Nanoparticles through Dual Solvent Exchange

We coated nanoparticles including iron oxide nanoparticles and quantum dots with phospholipid–PEG using the newly developed dual solvent exchange method and demonstrated that, compared with the conventional film hydration method, the coating efficiency and quality of coated nanoparticles can be significantly improved. A better control of surface coating density and the amount of reactive groups on nanoparticle surface is achieved, allowing conjugation of different moieties with desirable surface concentrations, thus facilitating biomedical applications of nanoparticles

Quantum Dot–Fluorescent Protein FRET Probes for Sensing Intracellular pH

Intracellular pH (pH_i) plays a critical role in the physiological and pathophysiological processes of cells, and fluorescence imaging using pH-sensitive indicators provides a powerful tool to assess the pH_i of intact cells and subcellular compartments. Here we describe a nanoparticle-based ratiometric pH sensor, comprising a bright and photostable semiconductor quantum dot (QD) and pH-sensitive fluorescent proteins (FPs), exhibiting dramatically improved sensitivity and photostability compared to BCECF, the most widely used fluorescent dye for pH imaging. We found that Förster resonance energy transfer between the QD and multiple FPs modulates the FP/QD emission ratio, exhibiting a >12-fold change between pH 6 and 8. The modularity of the probe enables customization to specific biological applications through genetic engineering of the FPs, as illustrated by the altered pH range of the probe through mutagenesis of the fluorescent protein. The QD-FP probes facilitate visualization of the acidification of endosomes in living cells following polyarginine-mediated uptake. These probes have the potential to enjoy a wide range of intracellular pH imaging applications that may not be feasible with fluorescent proteins or organic fluorophores alone

Substrate Stiffness Regulates Cellular Uptake of Nanoparticles

Nanoparticle (NP)-bioconjugates hold great promise for more sensitive disease diagnosis and more effective anticancer drug delivery compared with existing approaches. A critical aspect in both applications is cellular internalization of NPs, which is influenced by NP properties and cell surface mechanics. Despite considerable progress in optimization of the NP-bioconjugates for improved targeting, the role of substrate stiffness on cellular uptake has not been investigated. Using polyacrylamide (PA) hydrogels as model substrates with tunable stiffness, we quantified the relationship between substrate stiffness and cellular uptake of fluorescent NPs by bovine aortic endothelial cells (BAECs). We found that a stiffer substrate results in a higher total cellular uptake on a per cell basis, but a lower uptake per unit membrane area. To obtain a mechanistic understanding of the cellular uptake behavior, we developed a thermodynamic model that predicts that membrane spreading area and cell membrane tension are two key factors controlling cellular uptake of NPs, both of which are modulated by substrate stiffness. Our experimental and modeling results not only open up new avenues for engineering NP-based cancer cell targets for more effective in vivo delivery but also contribute an example of how the physical environment dictates cellular behavior and function

ZnS/Silica Nanocable Field Effect Transistors as Biological and Chemical Nanosensors

Compound semiconductor/isolator (ZnS/silica) core/shell nanocables have been used to fabricate single nanowire-based field effect transistors. Using the surface-adsorbed charged molecules as the gate, the nanocable-based devices show potential for label-free, real-time, and sensitive detection of biological species. After chemical modification, amine- and oxide-functionalized nanocables exhibit linear pH-dependent conductance, which could be elucidated in terms of the changes of surface charge during protonation and deprotonation. Selective biological recognition of nanocable sensors has been demonstrated using biotinylation

Magnetic Flap-Type Difunctional Sensor for Detecting Pneumatic Flow and Liquid Level Based on Triboelectric Nanogenerator

In recent years, the triboelectric nanogenerator (TENG) has attracted increasing attention because it not only converts various mechanical energy into electrical energy but also produces electrical signals as responses. On the basis of the TENG, a magnetic flap type difunctional sensor (MFTDS) has been developed to detect pneumatic flow and liquid level. Consisting of an outer magnetic flap, an inner magnetic float, and a conical cavity, its working mechanism and output characteristics were studied. The MFTDS detects pneumatic flows from 10 to 200 L/min with a flow resolution of 2 L/min. Compared with a commercial flow switch, the MFTDS results are in good agreement. Moreover, the MFTDS detects changes in liquid levels. The effects of liquid level height and flow rate on the performance of the MFTDS were measured and compared with a commercial liquid-level sensor. The results indicate that the output voltage of the MFTDS varies linearly with height but is independent of flow rate. The heights of liquid level from 30 to 130 mm were effectively detected. This work promotes the prospect for multifunctional triboelectric sensors

Surface Ligand Effects on Metal-Affinity Coordination to Quantum Dots: Implications for Nanoprobe Self-Assembly

The conjugation of biomolecules such as proteins and peptides to semiconductor quantum dots (QD) is a critical step in the development of QD-based imaging probes and nanocarriers. Such protein−QD assemblies can have a wide range of biological applications including in vitro protein assays and live-cell fluorescence imaging. One conjugation scheme that has a number of advantages is the self-assembly of biomolecules on a QD surface via polyhistidine coordination. This approach has been demonstrated using QDs that have different coating types, resulting in different interactions between the biomolecule and QD surface. Here, we report the use of a fluorescence resonance energy transfer (FRET) assay to evaluate the self-assembly of fluorescent proteins on the surface of QDs with eight distinct coatings, including several used in commercial preparations. The results of this systematic comparison can provide a basis for rational design of self-assembled biomolecule−QD complexes for biomedical applications