Dextran-Functionalized Super-nanoparticle Assemblies of Quantum Dots for Enhanced Cellular Immunolabeling and Imaging.

Colloidal semiconductor quantum dots (QDs) are a popular material for applications in bioanalysis and imaging. Although individual QDs are bright, some applications benefit from the use of even brighter materials. One approach to achieve higher brightness is to form super-nanoparticle (super-NP) assemblies of many QDs. Here, we present the preparation, characterization, and utility of dextran-functionalized super-NP assemblies of QDs. Amphiphilic dextran was synthesized and used to encapsulate many hydrophobic QDs via a simple emulsion-based method. The resulting super-NP assemblies or "super-QDs" had hydrodynamic diameters of ca. 90-160 nm, were characterized at the ensemble and single-particle levels, had orders-of-magnitude superior brightness compared to individual QDs, and were non-blinking. Additionally, binary mixtures of red, green, and blue (RGB) colors of QDs were used to prepare super-QDs, including colors difficult to obtain from individual QDs (e.g., magenta). Tetrameric antibody complexes (TACs) enabled simple antibody conjugation for selective cellular immunolabeling and imaging with both an epifluorescence microscope and a smartphone-based platform. The technical limitations of the latter platform were overcome by the increased per-particle brightness of the super-QDs, and the super-QDs outperformed individual QDs in both cases. Overall, the super-QDs are a very promising material for bioanalysis and imaging applications where brightness is paramount.

Fully Self-Assembled Silica Nanoparticle-Semiconductor Quantum Dot Supra-Nanoparticles and Immunoconjugates for Enhanced Cellular Imaging by Microscopy and Smartphone Camera.

There is a growing need for brighter luminescent materials to improve the detection and imaging of biomarkers. Relevant contexts include low-abundance biomarkers and technology-limited applications, where an example of the latter is the emerging use of smartphones and other nonoptimal but low-cost and portable devices for point-of-care diagnostics. One approach to achieving brighter luminescent materials is incorporating multiple copies of a luminescent material into a larger supra-nanoparticle (supra-NP) assembly. Here, we present a facile method for the preparation and immunoconjugation of supra-NP assemblies (SiO2@QDs) that comprised many quantum dots (QDs) around a central silica nanoparticle (SiO2 NP). The assembly was entirely driven by spontaneous affinity interactions between the constituent materials, which included imidazoline-functionalized silica nanoparticles, ligand-coated QDs, imidazole-functionalized dextran, and tetrameric antibody complexes (TACs). The physical and optical properties of the SiO2@QDs were characterized at both the ensemble and single-particle levels. Notably, the optical properties of the QDs were preserved upon assembly into supra-NPs, and single SiO2@QDs were approximately an order of magnitude brighter than single QDs and nonblinking. In proof-of-concept applications, including selective immunolabeling of breast cancer cells, the SiO2@QDs provided higher sensitivity and superior signal-to-background ratios whether using research-grade fluorescence microscopy or smartphone-based imaging. Overall, the SiO2@QDs are promising materials for enhanced bioanalysis and imaging.

Dextran-Functionalized Quantum Dot Immunoconjugates for Cellular Imaging.

Brightly luminescent semiconductor quantum dots (QDs) are ideal materials for cellular imaging and analysis because of their advantageous optical properties and surface area that supports multivalent conjugation of biomolecules. An important design consideration for effective use of these materials is a hydrophilic, biocompatible surface chemistry that provides colloidal stability and minimizes nonspecific interactions with biological molecules and systems. Dextran coatings are able to satisfy these criteria. Despite frequent use of dextran coatings with other nanomaterials (e.g., iron oxide nanoparticles), there has been little development and application of dextran coatings for QDs. In this chapter, we describe methods for the synthesis and characterization of a dextran ligand for QDs, including preparation of an immunoconjugate via tetrameric antibody complexes (TAC). The utility of these immunoconjugates is demonstrated through immunofluorescent labeling and imaging of overexpressed human epidermal growth factor receptor 2 (HER2) on the surface of SK-BR3 breast cancer cells.

Dextran Functionalization of Semiconducting Polymer Dots and Conjugation with Tetrameric Antibody Complexes for Bioanalysis and Imaging.

Brightly fluorescent semiconducting polymer dots (Pdots) are emerging as very useful probes for bioanalysis and imaging. Unfortunately, Pdot materials often suffer from limitations such as poor colloidal and physical stability, nonspecific adsorption, and relatively few reported surface chemistries and bioconjugate chemistries. To help address these limitations, we have developed dextran-functionalized Pdots (Dex-Pdots). This functionalization improves particle stability over a range of pH and at high ionic strength, hinders surface-induced unfolding, and enables the preparation of immunoconjugates via tetrameric antibody complexes (TAC). The utility of TAC-conjugated Dex-Pdots is demonstrated through a proof-of-concept fluorescence-linked immunosorbent assay (FLISA) for human erythropoietin (EPO), and through immunolabeling of human epidermal growth factor receptor 2 (HER2)-positive SK-BR3 breast cancer cells. The conjugates exhibited less nonspecific binding and greater specific binding than Pdots without dextran functionalization. Overall, dextran functionalization is a highly promising surface chemistry for biological applications of Pdots.