Yadong Xue

Quantum dots based electrochemiluminescent immunosensor by coupling enzymatic amplification with self-produced coreactant from oxygen reduction.

A highly sensitive competitive immunosensor based on the electrochemiluminescence (ECL) of quantum dots (QDs) was proposed by coupling with an enzymatic amplification. The fabrication process of the immunosensor was traced with atomic force microscopic images and electrochemical impedance spectra. The strong cathodic ECL emission of the immobilized QDs could be detected at a relatively low emission potential. The reduction of dissolved oxygen during the cathodic process provided a self-produced coreactant, H(2)O(2), for the ECL emission. Using human IgG (HIgG) as a model protein, upon the immuno-recognition of the immobilized HIgG to its antibody labeled simply with horseradish peroxidase, the ECL intensity decreased due to the steric hindrance of the proteins to electron transfer. The decrease could be greatly amplified by an enzymatic cycle to consume the self-produced coreactant, leading to a wide calibration range of 0.05 ng mL(-1) approximately 5 microg mL(-1) and a low limit of detection for the competitive immunoassay of HIgG. This immunosensor showed good stability and fabrication reproducibility. The immunoassays of practical samples showed acceptable results. This facile immunosensing strategy opened a new avenue for detection of proteins and application of QDs in ECL biosensing.

Highly sensitive electrocatalytic biosensing of hypoxanthine based on functionalization of graphene sheets with water-soluble conducting graft copolymer.

A novel electrocatalytic biosensing platform was designed by the functionalization of reduced graphene oxide sheets (RGO) with conducting polypyrrole graft copolymer, poly(styrenesulfonic acid-g-pyrrole) (PSSA-g-PPY), via π-π noncovalent interaction. The resulting nanocomposite could well disperse in water for at least 2 months with a solubility of 3.0 mg mL(-1). The nanocomposite was characterized with atomic force microscopy, X-ray photoelectron spectroscopy, ultraviolet-visible absorption, contact angle measurement, and electrochemical impedance spectroscopy. Based on the advantageous functions of PSSA-g-PPY and RGO, the functional nanocomposite modified platinum electrode showed high electrocatalytic activity toward the oxidation of hydrogen peroxide and uric acid in neutral media. Further, a hypoxanthine biosensor was constructed by combining the modified electrode with the enzymatic reaction of xanthine oxidase. The biosensor exhibited a wide linear response ranging from 3.0×10(-8) to 2.8×10(-5) M with a high sensitivity of 673±4 μA M(-1) cm(-2). The detection limit of 10nM at a signal-to-noise ratio of 3 was one order of magnitude lower than that reported previously. The assay results of hypoxanthine in fish samples were in a good agreement with the reference values. The water-soluble conducting copolymer could serve as an efficient species for functionalization and solubilization of graphene sheets in biosensing and biocatalytic applications.

In situ scanometric assay of cell surface carbohydrate by glyconanoparticle-aggregation-regulated silver enhancement.

A convenient and label-free scanometric approach for in situ cell surface carbohydrate assay was designed by integrating the bioconjugation and aggregation of glyconanoparticles, silver signal amplification, and spot test. The novel glyconanoparticles were prepared by a one-pot procedure. In the presence of lectin, using concanavalin A and mannose as a couple of model, the glyconanoparticles exhibited fast aggregation. The aggregation process could be inhibited by the specific recognition of lectin by the carbohydrate on the cell surface. Combining the gold nanoparticle catalyzed silver enhancement and scanometric detection, the number of cell surface carbohydrate groups could be conveniently read out. The average number of mannose units on a single living intact BGC cell was detected to be (4.5 +/- 0.4) x 10(7). This largely noninstrumental method took the advantages of a nanoparticle-based recognition and an aggregation-regulated signal amplification and avoided cell pretreatment and labeling processes. It could determine cell surface densities of different carbohydrates in parallel and thus would contribute considerably to meeting the challenges in decipherment of the glycomic codes.

Noncovalent functionalization of carbon nanotubes with lectin for label-free dynamic monitoring of cell-surface glycan expression

A kind of concanavalin A functionalized multiwalled carbon nanotube (ConA-MWCNT) was constructed by noncovalent assembly of ConA on carboxylated MWCNT with poly(diallyldimethylammonium) as a linker. The novel nanomaterial was characterized with scanning electron microscopy and atomic force microscopy. It incorporated both the specific recognition ability of lectin for cell-surface mannosyl groups and the unique electronic and mechanical properties of MWCNT. An electrochemical label-free method for cytosensing was proposed by constructing a ConA-MWCNT interface on a glassy carbon electrode, which showed a linear response to K562 cells ranging from 1 × 10(4) to 1 × 10(7) cellsmL(-1). The ConA-MWCNT interface could be further used for monitoring of dynamic variation of glycan expression on K562 cells in response to drugs. A facile and high-throughput optical method for the analysis of dynamic glycan expression on living cells was also developed by constructing an array of ConA-MWCNT spots on a glass slide. This method showed acceptable rapidity and low cost. The noncovalent functionalization of MWCNTs with lectins could be potentially applied in cell biological studies based on cell-surface glycan expression.

A simple electrochemical lectin-probe for in situ homogeneous cytosensing and facile evaluation of cell surface glycan.

This work constructed a novel electrochemical lectin-probe, ferrocene-concanavalin A (Fc-ConA), for in situ monitoring of cell surface glycan by incorporating the specific recognition ability of lectin to glycan and favorable electrochemical property of ferrocenyl group. The covalent conjugation of ConA with ferrocenyl group was achieved by a carbodiimide coupling reaction and proved with UV-vis absorption spectroscopy and infrared spectroscopy. Cyclic voltammetric behavior of Fc-ConA at glassy carbon electrode demonstrated a reversible diffusion-controlled process. A facile homogeneous cytosensing strategy was then developed using Fc-ConA probe for detection of K562 cells. The suspending cells specifically captured Fc-ConA via membrane mannosyl groups and decreased the concentration of free Fc-ConA, producing a response correlative with cell number and the content of cell surface glycan. A wide linear response to cells ranging from 1 × 10(4) to 1 × 10(7) cells mL(-1) with a calculated detection limit of 3000 cells mL(-1) was obtained. The lectin-probe could be conveniently used to in situ evaluate cell surface glycan. The average number of mannose moieties on single living K562 cell was detected to be 3.0 × 10(10), while this value increased by 81% on drug-treated cells. These results agreed with those from flow cytometric detection. This strategy presented a promising platform for homogeneous sensitive cytosensing and facile monitoring of carbohydrate expression on living cells in response to drugs.

In Situ Electrochemical Imaging of Membrane Glycan Expression on Micropatterned Adherent Single Cells

A scanning electrochemical microscopic (SECM) method for in situ imaging of four types of membrane glycan motifs on single adherent cells was proposed using BGC-823 human gastric carcinoma (BGC) cells as the model. These adherent cells were first micropatterned in the microwell of poly(dimethylsiloxane) membrane for precisely controlling the localized surface interaction, and the membrane glycans were then specifically recognized with corresponding lectins labeled with horseradish peroxidase (HRP). On the basis of the enzymatic oxidization of ferrocenylmethanol (FMA) by H(2)O(2) to yield FMA(+), the glycan expression level was detected by the reduction current of FMA(+) at the SECM tip. The cell-surface glycans could, thus, be in situ imaged by SECM at a single-cell level without peeling the cells from culture dish. Under the optimized conditions, four types of membrane glycan motifs showed statistically distinguishable expression levels. The SECM results for different glycan motifs on adherent single cells were consistent with those estimated by flow cytometric assay. This work provides a reliable approach for in situ evaluation of the characteristic glycopattern of single living cells and can be applied in cell biologic study based on cell surface carbohydrate expression.