Enhanced optical property of Au coated polystyrene beads for multi-color quantum dots encoding.

A novel quantum dots (QDs) fluorescent encoding method was demonstrated in this paper by using Au coated polystyrene (Au @ PS) beads. In the experiments, Au nanoparticles were deposited onto the polystyrene bead to form a stable Au coating through Layer-by-Layer assembly, and the surface morphology of the Au @ PS beads was studied by Scanning Electronic Microscope (SEM). Furthermore, the QDs encoding abilities, including the loading of QDs, the anti-photo bleaching ability and the multi-color encoding feasibility were studied using Au @ PS beads. The QDs leakage from doped QDs encoded Au @ PS was also studied. This study shows that Au particles improve the QDs encoding performance and the QD-encoded Au @ PS beads are the ideal material for the optical encoding.

Fluorophore-assisted carbohydrate electrophoresis as detection method for carbohydrate-protein interactions.

Fluorophore-assisted carbohydrate electrophoresis (FACE) is a straightforward, sensitive method for determining the presence and relative abundance of individual (oligo)saccharide in a(n) (oligo)saccharide mixture. The single terminal aldehydes of (oligo)saccharides were tagged with the charged fluorophore 8-aminonaphthalene-1,3,6-trisulfonate (ANTS), and separated with high resolution on the basis of size by polyacrylamide gel electrophoresis. ANTS fluorescence labeling is not biased by (oligo)saccharide length. Therefore, band fluorescence intensity is directly related to the relative abundance of individual (oligo)saccharide moieties in heterogeneous sample. In the same time, it also indicates that FACE can be used to investigate the interactions of carbohydrates and proteins.

Preparation of Au coated polystyrene beads and their application in an immunoassay.

A novel immunoassay method based on polystyrene beads coated with Au nanoparticles (Au@PS) is described. Au nanoparticles were prepared by reductive reaction, and then deposited on the surface of polystyrene beads to form Au coatings. Results indicated that the Au coatings had good stability and that human IgG was immobilized at a concentration of 16 microg/g Au@PS. FITC-labeled rabbit-anti-human IgG and FITC-labeled rabbit-anti-goat IgG were employed to react with the human IgG on Au@PS. Fluorescence imaging results showed that the reaction had good immuno-specificity. In addition, further experiments at the single-bead level indicated that the linear range was 0.05-15 microg/ml, and that the FITC signal could be detected even when the target antibody concentration was as low as 0.01 microg/ml. The assay results were compared with an enzyme-linked immunosorbent assay (ELISA), and showed relatively good reliability.

Quantum dot optical encoded polystyrene beads for DNA detection.

A novel multiplex analysis technology based on quantum dot (QD) optical encoded beads was studied. Carboxyl functionalized polystyrene beads, about 100 microm in size, were precisely encoded by the various ratios of two types of QDs whose emission wavelengths are 576 and 628 nm, respectively. Then the different encoded beads were covalently immobilized with different probes in the existing of sulfo-NHS and 1-[3-(Dimethylamino) propyl]-3-ethylcarbodiimide methiodide, and the probe density could reach to 3.1 mmol/g. These probe-linked encoded beads were used to detect the target DNA sequences in complex DNA solution by hybridization. Hybridization was visualized using fluorescein isothiocynate-labeled DNA sequences. The results show that the QDs and target signals can be obviously identified from a single-bead-level spectrum. This technology can detect DNA targets effectively with a detection limit of 0.2 microg/mL in complex solution.

Preparation of silica encapsulated quantum dot encoded beads for multiplex assay and its properties.

Novel -COOH modified polystyrene beads were prepared by sulfonation grafting, and the surface area and pore volume are greatly improved in comparison with the swelling-treated beads. The optimization coating time is 4 h, and the corresponding -COOH content is approximately 2.1 mmol/g. The scanning electron microscope results show that the silica particles deposited on the beads and formed a silica shell that decreases the leakage of quantum dots (QDs) preferably and improves the bar code stability greatly. The anti-photobleaching of silica-coated beads was studied systemically, and the results show that the half-decay time (t1/2) of the coated beads increases to 537 s--seven times longer than that of the uncoated ones. Further DNA probe hybridization experiments indicated that the coding signal and target signal can be detected simultaneously and that the assays based on these probe-conjugated silica/QD/polystyrene beads have good specificity and sensitivity that can detect a concentration as low as 0.01 microg/ml target DNA in denatured calf thymus DNA solution, indicating that it is feasible to use this kind of bead for multiplex analysis.

Synthesis, immunological activities, and scavenging ability toward superoxide anion of (1-->3)-beta-D-pentaglucoside and its epoxyalkyl derivatives.

The epoxyalkyl (1-->3)-beta-D-pentaglucosides 2 and 3 were synthesized in order by acetylation, glycosidation, oxidation, and deacetylation of 1. The immunological activities (superoxide anion production activity, phagocytic activity, and lymphocyte proliferation) and scavenging ability toward superoxide anion of (1-->3)-beta-D-pentaglucoside (1) and its epoxyalkyl derivatives (2 and 3) were investigated. Superoxide anion released from human blood monocytes was measured by the reduction of ferricytochrome c. Phagocytosis by peritoneal macrophages was detected through a teal ingesting that measured the chicken red blood cells (CRBC). Lymphocyte proliferation was determined by the MTT method. The scavenging ability of 1, 2, and 3 toward superoxide anions was evaluated by means of chemiluminescence (CL). The results showed that 2 and 3 had a little higher immunological activity and scavenging ability toward superoxide anion than 1, which indicated that the reducing end of the oligoglucosides was quite important for maximum biological activity.

The application of quantum dots as fluorescent label to glycoarray.

A sensitive, specific, and rapid method for the detection of carbohydrate-protein interactions was demonstrated using quantum dots (QDs) as a fluorescence label coupled with protein. 1,3-Dipolar cycloaddition between azide and alkyne was exploited to attach alpha-d-glucopyranoside to a C(14) hydrocarbon chain that noncovalently binds to the microtiter well surface, and the product formation was detected by both electrospray ionization-mass spectrometry (ESI-MS) and QD- (or fluorescein isothiocyanate (FITC))-conjugated lectin binding. It indicated that the peak intensity of the fluorescence emission was proportional to the initial concanavalin A (Con A) concentration in the range of 2 x 10(-3) micromol/L to 2 x 10(-2)mmol/L with a detection limit at least 100 times lower than that of the FITC-based method.

Synthesis of (R)-2,3-epoxypropyl(1-->3)-beta-D-pentaglucoside.

The title pentasaccharide was synthesized via a 2+3 strategy. The disaccharide donor, 3-O-acetyl-2-O-benzoyl-4,6-O-benzylidene-beta-D-glucopyranosyl-(1-->3)-2-O-benzoyl-4,6-O-benzylidene-alpha-D-glucopyranosyl trichloroacetimidate (8), was obtained by selective coupling of allyl 2-O-benzoyl-4,6-O-benzylidene-alpha-D-glucopyranoside with 3-O-acetyl-2-O-benzoyl-4,6-O-benzylidene-alpha-D-glucopyranosyl trichloroacetimidate (4), followed by deallylation, and trichloroacetimidation. Meanwhile, the trisaccharide acceptor, allyl 2-O-benzoyl-4,6-O-benzylidene-beta-D-glucopyranosyl-(1-->3)-2-O-benzoyl-4,6-O-benzylidene-beta-D-glucopyranosyl-(1-->3)-2-O-benzoyl-4,6-O-benzylidene-beta-D-glucopyranoside (12), was prepared by coupling of allyl 2-O-benzoyl-4,6-O-benzylidene-beta-D-glucopyranosyl-(1-->3)-2-O-benzoyl-4,6-O-benzylidene-beta-D-glucopyranoside with 4, followed by deacetylation. Condensation of 8 with 12, followed by epoxidation, and deprotection, gave the target pentaoside.

Synthesis, (1-->3)-beta-D-glucanase-binding ability and phytoalexin-elicitor activity of (R)-2,3-epoxypropyl (1-->3)-beta-D-pentaglucoside.

The (1-->3)-beta-D-pentaglucoside was synthesized as its (R)-2,3-epoxypropyl glycoside via 2+3 strategy. The disaccharide donor 8 was obtained by 3-selective coupling of 2 with 4, followed by deallylation, and trichloroacetimidation. Meanwhile, the trisaccharide acceptor 12 was prepared by coupling of 10 with 4, followed by deacetylation. Condensation of 8 with 12, followed by epoxidation, and deprotection, gave the target pentaoside. The results of these bioassays demonstrated that the (1-->3)-beta-D-glucanase was obviously inactivated by 15 with k(app)=3.79 x 10(-4) min(-1). At the same time, we found that the 15 was more active as compared to the laminaripentaose in eliciting phytoalexin accumulation in tobacco cotyledon tissue, and it could be kept longer time than laminaripentaose, which indicated it is much more stable than laminaripentaose.

Synthesis, protein-binding ability and phytoalexin-elicitor activity of epoxyalkyl (1-->3)-beta-D-oligoglucosides.

We describe a approach for the synthesis of (1-->3)-beta-D-oligosaccharide derivatives 10-18. 1-9 were synthesized by treating peracetylated (1-->3)-beta-D-oligosaccharides with the corresponding alkenyl alcohols and Lewis acid (SnCl(4)) catalyst. Epoxidation of the corresponding alkenyl oligoglucosides took place by m-CPBA. NaOMe in dry methanol was used for the deacetylation of the blocked derivatives, to give 10-18 in an overall yields of 25-32%. In subsequent glucan-binding protein of soybean assays, we found that 16 was most active, with an IC(50) value of 9 mM. However, the activities of 17, 18, 13, 14, 15, 10, 11, and 12 were gradually decreased. At the same time, we found 16 was most active as compared to the other (1-->3)-beta-D- oligoglucoside derivatives in eliciting phytoalexin accumulation in soybean cotyledon tissue, and 16 was kept longer time than (1-->3)-beta-D-glucohexaose, which indicated 16 is much more stable than (1-->3)-beta-D-glucohexaose.

Synthesis, (1-->3)-beta-D-glucanase-binding ability, and phytoalexin-elicitor activity of a mixture of 3,4-epoxybutyl (1-->3)-beta-D-oligoglucosides.

We describe a approach for the synthesis of a mixture of 3,4-epoxybutyl (1-->3)-beta-D-oligoglucosides. The particular (1-->3)-beta-D-glucan isolated from the cell walls of Saccharomyces cerevisiae was recovered from the aqueous medium as water-insoluble particles by the spray drying (GS) method, and it was characterized by FTIR spectroscopy. The acid-solubilized (1-->3)-beta-D-oligoglucosides were prepared by partial acid hydrolysis of glucan particles, which were qualitatively analyzed by fluorophore-assisted carbohydrate electrophoresis (FACE). The peracetylated 3-butenyl (1-->3)-beta-D-oligoglucosides were synthesized by treating peracetylated (1-->3)-beta-D-oligoglucosides with the 3-butenyl alcohols and a Lewis acid (SnCl4) catalyst. Epoxidation of the peracetylated 3-butenyl oligoglucosides took place with m-chloroperoxybenzoic acid (m-CPBA). NaOMe in dry methanol was used for the deacetylation of the blocked derivatives, to give the 3,4-epoxybutyl (1-->3)-beta-D-oligoglucoside mixture in an overall yield of 21%. The sample was analyzed by positive-ion electrospray ionization mass spectrometry (ESIMS). In a 3,4-epoxybutyl (1-->3)-beta-D-oligoglucoside-binding (1-->3)-beta-D-glucanase assay, we found that the (1-->3)-beta-D-glucanase was obviously inactivated by the 3,4-epoxybutyl (1-->3)-beta-D-oligoglucosides. At the same time, we found the 3,4-epoxybutyl (1-->3)-beta-D-oligoglucoside mixture was more active as compared to the underivatized oligoglucoside mixture in eliciting phytoalexin accumulation in tobacco cotyledon tissue. Furthermore, it could be kept for a longer time than a (1-->3)-beta-D-oligoglucoside mixture, which indicated it is much more stable than (1-->3)-beta-D-oligoglucosides.