Immunoassay by capillary electrophoresis with quantum dots

The application of quantum dots in capillary electrophoresis immunoassay was studied for the first time. Quantum dots were conjugated with antibody and subsequently tested by electrophoretic separation of free antibody and antibody-antigen complex. Antibody was fluorescently labeled by quantum dots via conjugation procedures and its electrophoretic characteristics were effectively modified due to the attachment of quantum dots. The determination of human IgM by direct CE based immunoassay could be easily achieved by simply changing the pH value of separation buffer. Polymer additive influenced the separation too but the effect was not as significant as buffer pH adjustment. Satisfactory separation of complex from free antibody could be achieved with 20mM sodium tetraborate as separation buffer, at pH 9.8. The immunoassay application of quantum dots in CE offers considerable advantages and can be readily applied to other large bio-molecules.     

Characterization of quantum dots using capillary zone electrophoresis

Commercially available quantum dots (QDs) were characterized using CE. The CE instruments were laboratory-built, each being capable of both electrokinetic and hydrodynamic injection. Modes of detection include UV absorption and LIF. The CE-LIF system was further modified to handle microliter sample volumes during injection. Sodium phosphate (5-25 mM, pH 7.5-11) was found to be a good buffer electrolyte. Sodium mercaptoproprionate CdTe/CdS (ADS620) QDs and carboxylic acid CdSe/ZnS (T2-Evitag) QDs yielded high separation efficiencies of N = 1.5x10(6) plates at t(M) = 10 min and N = 1.0x10(5) plates at t(M) = 3.8 min, respectively. Apparently the EDC/sulfo-NHS bioconjugation chemistry worked well with the neutral T2-Evitag QDs, but not so well with the negatively charged ADS620 QDs. This preliminary knowledge will serve as a basis for new CE immunoassay studies of QD-biomolecule conjugates and their immunocomplexes with target analytes.     

Characterization and separation of semiconductor quantum dots and their conjugates by capillary electrophoresis

Semiconductor quantum dots (QDs) are very important luminescent nanomaterials with a wide range of potential applications. Currently, QDs as labeling probes are broadly used in bioassays, including immunoassay, DNA hybridization, and bioimaging, due to their excellent physical and chemical properties, such as broad excitation spectra, narrow and size‐dependent emission profiles, long fluorescence life time, and good photostability. The characterization of QDs and their conjugates is crucial for their wide bioapplications. CE has become a powerful tool for the separation and characterization of QDs and their conjugates. In this review, some CE separation models of QDs are first introduced, mainly including CZE, CGE, MEKC, and ITP. And then, some key applications, such as the measurements of size, surface charge, and concentration of QDs and the characterization of QDs conjugates (e.g. QD–protein, QD–DNA, QD–small molecule), are also described. Finally, future perspectives are discussed.     

Hydrophilization of CdSe quantum dots with surfactants

A procedure has been developed for hydrophilization of CdSe semiconductor quantum dots with anionic, cationic, and nonionic surfactants. Depending on the type of a surfactant, these particles may carry positive, negative, or no charges. In spite of the universality of the proposed procedure, the hydrophilization with specific surfactants must be carried out at specific concentrations of reagents and medium pH values. Under optimum conditions, the transfer of the quantum dots to water substantially increases the quantum yield of their luminescence. Conditions have been found under which the luminescence properties of the hydrophilic particles remain almost unchanged for a long time period.     

Characterization of CdSe nanocrystals coated with amphiphiles. A capillary electrophoresis study

We have synthesized CdSe nanocrystals (NCs) possessing a trioctylphosphine surface passivation layer and modified with amphiphilic molecules to form a surface bilayer. The NCs covered with single amphiphiles are not stable in aqueous solution, but a mixed amphiphilic system is shown to provide stability in solution over several months. The solutions of the modified NCs were characterized by UV-Vis absorbance, photoluminescence, and transmission electron microscopy. An electrophoretic study revealed two operational modes. The first relies on the enrichment of NCs using a micellar plug as a tool. The accumulation of NCs at the plug-electrolyte buffer interface results in a sharp peak. By controlling the electrophoretic conditions, nanocrystals were forced to exit a micellar plug into an electrolyte buffer. We conclude that a system consisting of modified nanocrystals and a micellar plug can act as a mixed pseudomicellar system, where modified nanocrystals play the role of pseudomicelles.FigureElectrophoretic focusing of amphiphile coated CdSe nanocrystals using a micellar plug. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1007/s00604-011-0727-8) contains supplementary material, which is available to authorized users.     

Characterization of quantum dot bioconjugates by capillary electrophoresis with laser-induced fluorescent detection

In this paper, we present a universal, highly efficient and sensitive method for the characterization of quantum dot (QD) bioconjugates based on capillary electrophoresis with laser-induced fluorescent (LIF) detection. We first prepared CdTe QDs in aqueous phase by a chemical route with mercaptopropionic acid as a ligand, and then were coupled to certain proteins using bifunctional linkage reagent or electrostatic attraction. The QD bioconjugates were characterized by capillary electrophoresis with LIF detection. We found that QD bioconjugates were efficiently separated with free QDs by the optimization of buffer pH. Furthermore, we found that ultrafiltration was an effective and simple approach to purify QD conjugates with bovine serum albumin (BSA). Due to their broad absorption spectra and size dependent emission wavelength tunability, QDs can be excited to emit different colour fluorescence using a single wavelength laser source, and therefore, we believe that CE with LIF detection will become a universal and efficient tool for the characterization of QD bioconjugates.     

Role of surface ligands in optical properties of colloidal CdSe/CdS quantum dots

In order to study the role of surface ligands in determining optical properties of colloidal quantum dots (QDs), we have selectively fabricated and studied CdSe/CdS core-shell QDs with strongly confined electron and hole states attached with commonly used surface ligands. Optical properties, viz. absorption and fluorescence of these QDs, are characterized from which salient changes have been observed for different ligand substitutions which, through theoretical analysis, can be associated with electronic structure properties of the QD-ligand composite systems, in particular localization of wave functions of electrons and holes in the QDs and the band matching of the HOMO-LUMO gap of the ligands. The findings can be utilized to facilitate the understanding and optimization of properties of QD biomarkers with functionalizing surface ligands for targeting cellular objects.     

Probing cytotoxicity of CdSe and CdSe/CdS quantum dots

Toxicities of CdSe and CdSe/CdS quantum dots(QDs) synthesized by ultrasound-assisted methods were investigated in vitro and in vivo.Five human cell lines were used to assess the cytotoxicity of as-prepared CdSe and CdSe/CdS by assays of MTT viability,red blood cell hemolysis,flow cytometry,and fluorescence imaging.The results show that these QDs may be cytotoxic by their influence in S and G2 phases in cell cycles.The cytotoxicity of QDs depends on both the physicochemical properties and related to target cells.     

Resolving quantum dots and peptide assembly and disassembly using bending capillary electrophoresis

Despite the numerous techniques developed for the studying nanoparticle and peptide interaction nowadays, sensitive and convenient assay in the process of flow, especially to simulate the self‐assembly of quantum dots (QDs) and peptide inflow in blood vessels, still remains big challenges. Here, we report a novel assay for studying the self‐assembly of QDs and peptide, based on CE using a bending capillary. We demonstrate that the semicircles numbers of the bending capillary affect the self‐assembly kinetics of CdSe/ZnS QDs and ATTO‐D₃LVPRGSGP₉G₂H₆ peptide. Moreover, benefitting from this novel assay, the effect of the position on the self‐assembly has also been realized. More importantly, we also demonstrate that this novel assay can be used for studying the stability of the QDs–peptide complex inflow. We believe that our novel assay proposed in this work could be further used as a general strategy for the studying nanoparticle–biomolecule interaction or biomolecule–biomolecule interaction.     

Capillary electrophoresis immunoassays with conjugated quantum dots

Water-soluble CdTe quantum dots (QDs) and their conjugates with antibodies and antigenes were prepared by optimized procedures for applications in CE immunoassays. The QD size of 3.5 nm, excitation spectrum in the range of 300-500 nm, the maximum wavelength of the emission spectrum at 610 nm, quantum yield of 0.25 and luminescence lifetimes in the range of 3.6-43 ns were determined. The 0.1 M solution of TRIS/TAPS (pH 8.3) was found to be the optimum buffer for the separation of the antiovalbumin-ovalbumin immunocomplex from the free conjugates of QDs.     

Characterization and 2D self-assembly of CdSe quantum dots at the air-water interface

Langmuir film properties, UV-vis spectroscopy, epifluorescence microscopy, and transmission electron microscopy were used to study CdSe quantum dots (QDs) in 2D. By combining these results, it was possible to determine the molar absorptivity, limiting nanoparticle area, luminescence property, and arrangement of the QDs in the monolayer films at the air-water interface. Either trioctylphosphine oxide (TOPO) or 1-octadecanethiol (ODT) stabilized the QDs. The data collected reveal that TOPO forms close-packed monolayers on the surface of the QDs and that ODT-stabilized QDs undergo alkyl chains interdigitation. It was also found that varying the nanoparticle size, nature of surfactant, surface pressure, and mixed monolayers could help engineer the 2D self-assembly of the QDs at the air-water interface. Of practical importance is the transfer of these monolayer films onto hydrophilic or hydrophobic solid substrates, which could be successfully accomplished via the Langmuir-Blodgett film deposition technique.     

CdSe quantum dots in a columnar matrix

Cadmium selenide quantum dots (QDs) have been dispersed in a discotic liquid crystal columnar matrix. Thermophysical properties of these liquid crystal nanocomposites were investigated by UV-Vis spectroscopy, photoluminescence spectroscopy, differential scanning calorimetry, polarizing optical microscopy, DC conductivity and small angle X-ray diffraction.     

Photoassisted synthesis of CdSe and core-shell CdSe/CdS quantum dots

This paper describes the synthesis of core-shell CdSe/CdS quantum dots (QDs) in aqueous solution by a simple photoassisted method. CdSe was prepared from cadmium nitrate and 1,1-dimethylselenourea precursors under illumination for up to 3 h using a pulsed Nd:YAG laser at 532 nm. The effects that the temperature and the laser irradiation process have on the synthesis of CdSe were monitored by a series of experiments using the precursors at a Cd:Se concentration ratio of 4. Upon increasing the temperature (80-140 degrees C), the size of the CdSe QDs increases and the time required for reaching a maximum photoluminescence (PL) is shortened. Although the as-prepared CdSe QDs possess greater quantum yields (up to 0.072%) compared to those obtained by microwave heating (0.016%), they still fluoresce only weakly. After passivation of CdSe (prepared at 80 degrees C) by CdS using thioacetamide as the S source (Se:S concentration ratio of 1) at 80 degrees C for 24 h, the quantum yield of the core-shell CdSe/CdS QDs at 603 nm is 2.4%. Under UV irradiation of CdSe/CdS for 24 h using a 100-W Hg-Xe lamp, the maximum quantum yield of the stable QDs is 60% at 589 nm. A small bandwidth (W1/2 < 35 nm) indicates the narrow size distribution of the as-prepared core-shell CdSe/CdS QDs. This simple photoassisted method also allows the preparation of differently sized (3.7-6.3-nm diameters) core-shell CdSe/CdS QDs that emit in a wide range (from green to red) when excited at 480 nm.     

Studies on bioconjugation of quantum dots using capillary electrophoresis and fluorescence correlation spectroscopy

In this paper, we systematically investigated the conjugation of quantum dots (QDs) with certain biomolecules using capillary electrophoresis (CE) and fluorescence correlation spectroscopy (FCS) methods. Commercial QDs and aqueous-synthesized QDs in our lab were used as labeling probes, certain bio-macromolecules, such as proteins, antibodies, and enzymes, were used as mode samples, and 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC) and N-hydroxysulfo-succinimide (Sulfo-NHS) were used as linking reagents. We studied the effects of certain factors such as the isoelectric points (pIs) of bio-macromolecules and buffer pH on the bioconjugation of QDs, and found that the pIs of bio-macromolecules played an important role in the conjugation reaction. By the optimization of the buffer pH some proteins with different pIs were efficiently conjugated with QDs using EDC and Sulfo-NHS as linking agents. Furthermore, we on-line investigated the kinetic process of QDs-bioconjugation by FCS and found that the conjugation reaction of QDs with protein was rapid and the reaction process almost completed within 10 min. We also observed that QDs conjugated with proteins were stable for at least 5 days in phosphate buffer. Our work described here will be very helpful for the improvement of the QDs conjugation efficiency in bioapplications.     

Electrochemical properties of CdSe and CdTe quantum dots

Semiconductor nanocrystal quantum dots (QDs), owing to their unique opto-electronic properties determined by quantum confinement effects, have been the subject of extensive investigations in different areas of science and technology in the past two decades. The electrochemical behaviour of QDs, particularly for CdSe and CdTe nanocrystals, has also been explored, although to a lesser extent compared to the optical properties. Voltammetric measurements can be used to probe the redox levels available for the nanocrystals, which is an invaluable piece of information if these systems are involved in electron transfer processes. Electrochemical data can also foster the interpretation of the spectroscopic properties of QDs, and give insightful information on their chemical composition, dimension, and surface properties. Hence, electrochemical methods constitute in principle an effective tool to probe the quality of QD samples in terms of purity, size dispersion, and surface defects. The scope of this critical review is to discuss the results of electrochemical studies carried out on CdSe and CdTe core and core-shell semiconductor nanocrystals of spherical shape. Examples of emerging or potential applications that exploit electroactive quantum dot-based systems will also be illustrated.     

Interactions between CdTe quantum dots and DNA revealed by capillary electrophoresis with laser‐induced fluorescence detection

Quantum dots (QDs) are one of the most promising nanomaterials, due to their size‐dependent characteristics as well as easily controllable size during the synthesis process. They are promising label material and their interaction with biomolecules is of great interest for science. In this study, CdTe QDs were synthesized under optimal conditions for 2 nm size. Characterization and verification of QDs synthesis procedure were done by fluorimetric method and with CE. Afterwards, QDs interaction with chicken genomic DNA and 500 bpDNA fragment was observed employing CE‐LIF and gel electrophoresis. Performed interaction relies on possible matching between size of QDs and major groove of the DNA, which is approximately 2.1 nm.     

Switching-on superparamagnetism in Mn/CdSe quantum dots

Mn ion doping of CdSe and other semimagnetic quantum dot (QDs) alloys has been an area of active speculation for over a decade. We report evidence of Mn(II) doping of CdSe grown from a cubic single source precursor that is superparamagnetic (SPM) with a blocking temperature of 40 K following thermal annealing. Prior to thermal annealing the 4 nm Mn/CdSe (1% Mn) QDs exhibit mainly paramagnetic behavior between 300 and 2 K, with a weak antiferromagnetic exchange. Following thermal annealing of the sample, high-temperature ferromagnetic exchange is observed in the magnetization data with the onset of an SPM phase at 40 K that exhibits a coercivity of 0.1 T at 2 K. The switching-on of SPM behavior is believed to be linked to ion migration with formation of (Se-Mn-Se-Mn-Se-Mn)n centers within the nanocrystal that exhibit coupled magnetic moments. Electron paramagnetic resonance (EPR) provides evidence of two distorted T(d) Mn core sites, a clustered site (dipolar broadened), and a localized Mn site (hyperfine-split). The ratio of the EPR signature for the dipolar broadened site increases following annealing and shows a hysteretic response around the blocking temperature. These observations suggest that thermal annealing results in enhanced cluster formation explaining the onset of the SPM phase in these nanoscale materials. Evidence of SPM behavior is evident in the field-dependent non-Langevin magnetization with a tangential loss in the ac-magnetic susceptibility and the Mydosh parameter (phi = 0.16).     

Characterization of synthetic polyelectrolytes by capillary electrophoresis

Capillary electrophoresis in entangled polymer solutions was applied to determine the molecular mass and polydispersity of polyelectrolytes. The separation selectivities of different polyethylene glycols as buffer additive can be correlated to their average molecular mass. A universal curve correlating the selectivity and the molecular mass could be obtained by using the instrinsic viscosity of the polyethylene glycol. The separation of poly(2-vinylpyridine) standards was compared to the separation of poly(4-vinylpyridine) standards. An indirect detection system was developed to characterize the cationic polyelectrolyte polydiallyldimethyl ammonium chloride. Various polymers with oppositely charged groups (polycarboxybetaines) were investigated with respect to structure dependence, pH dependence and molecular mass dependence of interand intramolecular association.     

Characterization of carboxylated nanolatexes by capillary electrophoresis

Poly(styrene-co-acrylic acid) (St/AA) and poly(styrene-co-methacrylic acid) (St/MA) nanolatexes with different acid contents were prepared by emulsion copolymerization and were analyzed by capillary electrophoresis (CE) and by laser doppler velocimetry (LDV). Due to the intrinsic differences in the methodologies, CE (separative technique) and LDV (zetametry, nonseparative technique) lead to very different electrophoretic mobility distributions. Beyond these differences, the variation of the electrophoretic mobility is a complex and nonlinear function of the hydrodynamic radius, the ionic strength, and the zeta potential. To gain better insight on the influence of the ionic strength and the acid content on the electrophoretic behavior of the nanolatexes, the electrophoretic mobility data were changed into surface charge densities using the O'Brien, White, and Ohshima modeling. This approach leads to the conclusion that the surface charge density is mainly controlled at high ionic strength (～50 mM) by the adsorption of anionic surfactants coming from the sample. On the contrary, at low ionic strength, and/or in the presence of neutral surfactant in the electrolyte, the acid content was the main parameter controlling the surface charge density of the nanolatexes.     

Characterization of humic substances by capillary electrophoresis

Capillary electrophoresis was used for the separation of humic acid (HA) from peat, soil, and vermicompost. The electropherograms show the presence of at least three peaks eluted between 6 and 11 min for all HA. The best analysis resolution was obtained with the use of borate buffers at pH 8.9. The HA analyzed have structural and charge similarity, which increases the difficulty of separation. Therefore, the shape of the peaks is broad and the CE profiles of all HA are similar. It is reasonable to assume that the broad band in the three regions is due to the acidic groups that have a similar structure. By comparing the results obtained for HA extracted from soil, peat, vermicompost, and the commercial sample, HA from peat had the major carbon content.