共查询到20条相似文献,搜索用时 46 毫秒
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Techniques that can effectively separate protein–peptide complexes from free peptides have shown great value in major histocompatibility complex (MHC)–peptide binding studies. However, most of the available techniques are limited to measuring the binding of a single peptide to an MHC molecule. As antigen presentation in vivo involves both endogenous ligands and exogenous antigens, the deconvolution of multiple binding events necessitates the implementation of a more powerful technique. Here we show that capillary electrophoresis coupled to fluorescence detection (CE–FL) can resolve multiple MHC–peptide binding events owing to its superior resolution and the ability to simultaneously monitor multiple emission channels. We utilized CE–FL to investigate competition and displacement of endogenous peptides by an immunogenic gluten peptide for binding to HLA-DQ2. Remarkably, this immunogenic peptide could displace CLIP peptides from the DQ2 binding site at neutral but not acidic pH. This unusual ability of the gluten peptide supports a direct loading mechanism of antigen presentation in extracellular environment, a property that could explain the antigenicity of dietary gluten in celiac disease. 相似文献
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LI ZhiYong ZHANG QianYun ZHAO LiXia LI ZhenJia HU GuoMao LIN Jin-Ming & WANG Shan State Key Laboratory of Environmental Chemistry Ecotoxicology Research Center for Eco-Environmental Sciences Chinese Academy of Sciences Beijing China 《中国科学:化学》2010,(4)
A micro-plate magnetic chemiluminescence immunoassay was developed for rapid and high throughput detection of carcinoembryonic antigens (CEA) in human sera. This method was based on a sandwich immunoreaction of fluorescein isothiocyanate (FITC)-labeled anti-CEA antibodies, CEA antigens, and horseradish peroxidase (HRP)-conjugated anti-CEA antibodies in mi- cro-plate. The immunomagnetic particles coated with anti-FITC antibodies were used as the solid phase for the immunoassay. The separation procedure was c... 相似文献
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Giaume D Poggi M Casanova D Mialon G Lahlil K Alexandrou A Gacoin T Boilot JP 《Langmuir : the ACS journal of surfaces and colloids》2008,24(19):11018-11026
Luminescent inorganic nanoparticles are now widely studied for their applications as biological probes for in vitro or in vivo experiments. The functionalization of the particles is a key step toward these applications, since it determines the control of the coupling between the particles and the biological species of interest. This paper is devoted to the case of rare earth doped oxide nanoparticles and their functionalization through their surface encapsulation with a functional polysiloxane shell. The first step of the process is the adsorption of silicate ions that will act as a primary layer for the further surface polymerization of the silane, either aminopropyltriethoxysilane (APTES) or glycidoxypropyltrimethoxysilane (GPTMS). The amino- or epoxy- functions born by the silane allow the versatile coupling of the particles with bio-organic species following the chemistry that is commonly used in biochips. Special attention is paid to the careful characterization of each step of the functionalization process, especially concerning the average number of organic functions that are available for the final coupling of the particles with proteins. The surface density of amino or epoxy functions was found to be 0.4 and 1.9 functions per square nanometer for GPTMS and APTES silanized particles, respectively. An example of application of the amino-functionalized particles is given for the coupling with alpha-bungarotoxins. The average number (up to 8) and the distribution of the number of proteins per particle are given, showing the potentialities of the functionalization process for the labeling of biological species. 相似文献
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Natural killer T cells (NKT cells) play a central role in regulating immune responses influencing conditions ranging from autoimmune to infectious diseases. NKT cell responses are induced by recognition of glycolipid antigens presented by CD1d, an antigen presentation protein. In the last 10 years great strides have been made in understanding the types of glycolipids recognized by NKT cells. These advances have included determination of the lipid and carbohydrate recognition requirements for stimulation and identification of "natural" antigens for these cells. 相似文献
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Patricia Pereira Aguilar Irene Gonzlez‐Domínguez Tobias Amadeus Schneider Francesc Gdia Laura Cervera Alois Jungbauer 《Journal of separation science》2019,42(16):2640-2649
At‐line static light scattering and fluorescence monitoring allows direct in‐process tracking of fluorescent virus‐like particles. We have demonstrated this by coupling at‐line multi‐angle light scattering and fluorescence detectors to the downstream processing of enveloped virus‐like particles. Since light scattering intensity is directly proportional to particle concentration, our strategy allowed a swift identification of product containing fractions and rapid process development. Virus‐like particles containing the Human Immunodeficiency Virus‐1 Gag protein fused to the Green Fluorescence protein were produced in Human Embryonic Kidney 293 cells by transient transfection. A single‐column anion‐exchange chromatography method was used for direct capture and purification. The majority of host‐cell protein impurities passed through the column without binding. Virus‐like particles bound to the column were eluted by linear or step salt gradients. Particles recovered in the step gradient purification were characterized by nanoparticle tracking analysis, size exclusion chromatography coupled to multi‐angle light scattering and fluorescence detectors and transmission electron microscopy. A total recovery of 66% for the fluorescent particles was obtained with a 50% yield in the main product peak. Virus‐like particles were concentrated 17‐fold to final a concentration of 4.45 × 1010 particles/mL. Simple buffers and operation make this process suitable for large scale purposes. 相似文献
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Yoshinori Sohma Ryoji Fujita Shigeo Katoh Eizo Sada 《Applied biochemistry and biotechnology》1993,38(3):179-188
In order to clarify the effects of the differences in physical states of antigens on recognition by antibodies in immunoassays,
the binding characteristics of an antipeptide polyclonal antibody to the peptide and the corresponding protein were studied.
The reactivity in the immunoliposome assay (ILA), as well as in the double-antibody sandwich ELISA, was identical to that
in the solution. These results indicate that the conformation of liposome-bound antigen is changed little by coupling to liposomes
and is almost the same as that of the native antigen in the liquid phase. It is desirable to assay by double-antibody sandwich
ELISA or ILA to detect native proteins, and the latter is very easily performed. 相似文献
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Simultaneous bioassays in a microfluidic channel on plugs of different magnetic particles 总被引:1,自引:0,他引:1
Magnetic particles coated with specific biomolecules are often used as solid supports for bioassays but conventional test tube based techniques are time consuming and labour intensive. An alternative is to work on magnetic particle plugs immobilised inside microfluidic channels. Most research so far has focussed on immobilising one type of particle to perform one type of assay. Here we demonstrate how several assays can be performed simultaneously by flushing a sample solution over several plugs of magnetic particles with different surface coatings. Within a microchannel, three plugs of magnetic particles were immobilised with external magnets. The particles featured surface coatings of glycine, streptavidin and protein A, respectively. Reagents were then flushed through the three plugs. Molecular binding occurred between matching antigens and antibodies in continuous flow and was detected by fluorescence. This first demonstration opens the door to a quicker and easier technique for simultaneous bioassays using magnetic particles. 相似文献
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Howard H. Weetall 《Applied biochemistry and biotechnology》1993,41(3):157-188
Enzymes were first immobilized on inorganic supports through silane coupling agents over 25 yr ago. Since that initial report, literally hundreds of laboratories have utilized this methodology for the immobilization of enzymes, antigens, antibodies, receptors, and other high and low mol wt compounds. Today silane coupling is one of the commonly used techniques in the arsenal of the biochemist for the binding of material of all sorts to inorganic surfaces. Inorganic materials come in a variety of shapes, sizes, and characteristics. Today silane coupling is one of the most used coupling methods for the preparation of biosensing devices. Sol-gel entrapped enzymes are also produced by the application of silane technology by the polymerization of the silane to form glass-like materials with entrapped protein. This review will discuss the general preparation and characterization of silane coupled proteins with special emphasis on enzymes and describe in detail the actual methods for the silanization and specific chemical coupling of proteins to the silanized carrier. 相似文献
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Proteasome function in antigen presentation: immunoproteasome complexes, Peptide production, and interactions with viral proteins 总被引:4,自引:0,他引:4
Proteasomes are the major nonlysosomal protein degradation machinery in eukaryotic cells and they are largely responsible for the processing of antigens for presentation by the MHC class I pathway. This review concentrates on recent developments in the area of antigen processing. Specialized proteasomes called immunoproteasomes and an 11S regulator of proteasomes (PA28) are induced by interferon-gamma, but it is not entirely clear why changes in proteasome structure are beneficial for antigen presentation. Different proteasome complexes have distinct subcellular distributions and subtle differences in cleavage specificity. Thus it is likely that the efficiency of production of MHC class I binding peptides varies in different locations. Immunoproteasome subunits are enriched at the ER where TAP transports peptides for association with newly synthesized MHC class I molecules. There is recent evidence to suggest that antigen presentation from viral expression vectors, or from peptides that are either delivered by bacterial toxins or derived from signal peptides, require proteasome activity for generation of the correct C-terminus of the epitope. The correct N-terminus may be generated by recently identified ER associated aminopeptidases. A number of viral protein interactions with proteasome subunits have been reported and such interactions may interfere with host anti-viral defenses and also contribute to mechanisms of cell transformation. 相似文献
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Yonghui Liu Mingjing Li Haomiao Zhu Zhe Jing Xiaona Yin Kun Wang Zhangyong Hong Wei Zhao 《中国化学快报》2021,32(6):1963-1966
We have developed a MUC1 antigen-based antitumor vaccine loaded on alum colloid encapsulated insideβ-glucan particles(GP-Al). The constructed vaccine induced strong MUC1 antigen specific Ig G antibody titers and enhanced CD8~+ T cells cytotoxic effect to kill tumor cells. These results indicated that GP-Al can be served as an efficient delivery system and adjuvant for the development of cancer vaccines especially small molecule antigens based cancer vaccines. 相似文献
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Matsunaga T Maeda Y Yoshino T Takeyama H Takahashi M Ginya H Aasahina J Tajima H 《Analytica chimica acta》2007,597(2):331-339
Magnetic beads have served as a conventional bioassay platform in biotechnology. In this study, a fully automated immunoassay was performed using novel nano- and microbead-composites constructed by assembling nano-magnetic beads onto polystyrene microbeads, designated ‘Beads on Beads’. Nano-sized bacterial magnetic particles (BacMPs) displaying the immunoglobulin G (IgG)-binding domain of protein A (ZZ domain) were used for the construction of ‘Beads on Beads’ via the interaction of biotin-streptavidin. The efficient assembly of ‘Beads on Beads’ was performed by gradual addition of biotin-labeled BacMPs onto streptavidin-coated polystyrene microbeads. Approximately 2000 BacMPs were uniformly assembled on a single microbead without aggregation. The constructed ‘Beads on Beads’ were magnetized and separated from the suspension by using an automated magnetic separation system with a higher efficiency than BacMPs alone. Furthermore, fully automated detection of prostate-specific antigens was performed with the detection limit of 1.48 ng mL−1. From this preliminary assay, it can be seen that ‘Beads on Beads’ could be a powerful tool in the development of high-throughput, fully automated multiplexed bioassays. 相似文献
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The homobifunctional crosslinker 1,4‐phenylene diisothiocyanate (PDC) was coupled to amine‐terminated magnetic particles, and human IgG, streptavidin, protein G and protein A were immobilized on the activated magnetic particles. The coupling of PDC to the amine‐terminated magnetic particles was completed in 10 min, and 1 mg of activated magnetic particles was able to immobilize 95 (g of protein G, 120 µg of protein A, 160 µg of streptavidin and 280 µg of IgG. Ultraviolet‐visible spectroscopy, FTIR spectroscopy and electron micrography were used to characterize the functional particles. The results indicated that PDC was successfully coupled to the surface of the amine‐terminated magnetic particles, and the proteins were effectively immobilized on their surface. The activity of protein G immobilized on the activated magnetic particles was confirmed by its ability to purify IgG from plasma. 相似文献
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Kaiyuan Ni Taokun Luo Guangxu Lan August Culbert Yang Song Tong Wu Xiaomin Jiang Wenbin Lin 《Angewandte Chemie (International ed. in English)》2020,59(3):1108-1112
Checkpoint blockade immunotherapy (CBI) awakes a host innate immune system and reactivates cytotoxic T cells to elicit durable response in some cancer patients. Now, a cationic nanoscale metal–organic framework, W‐TBP, is used to facilitate tumor antigen presentation by enabling immunogenic photodynamic therapy (PDT) and promoting the maturation of dendritic cells (DCs). Comprised of dinuclear WVI secondary building units and photosensitizing 5,10,15,20‐tetra(p‐benzoato)porphyrin (TBP) ligands, cationic W‐TBP mediates PDT to release tumor associated antigens and delivers immunostimulatory CpG oligodeoxynucleotides to DCs. The enhanced antigen presentation synergizes with CBI to expand and reinvigorate cytotoxic T cells, leading to superb anticancer efficacy and robust abscopal effects with >97 % tumor regression in a bilateral breast cancer model. 相似文献
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A novel nanocomposite based on TiO2–protein for functional applications was prepared and characterized. The composite was made by covalent immobilisation of bovine serum albumin (BSA) on to the TiO2 particles. BSA was attached to the TiO2 powder through a three-step process. This involves the functionalization of the TiO2 particles with silane and the subsequent coupling by glutaraldehyde to free NH2 groups of the protein. Formation of Schiff's base was confirmed by IR spectroscopy and the optimum loading of the BSA was found to be 73.61%. Surface morphology of the composite was studied by SEM and TEM. Thermal analysis of TiO2–BSA composite was carried out by TGA and DSC. Structural variation of the BSA after immobilisation was studied by CD spectra. 相似文献
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《化学:亚洲杂志》2017,12(1):159-167
Functional pairing of cellular glycoconjugates with tissue lectins is a highly selective process, whose determinative factors have not yet been fully delineated. Glycan structure and modes of presentation, that is, its position and density, can contribute to binding, as different members of a lectin family can regulate degrees of responsiveness to these factors. Using a peptide repeat sequence motif of the glycoprotein mucin‐1, the principle of introducing synthetic (glyco)peptides with distinct variations in these three parameters to an array‐based screening of tissue lectins is illustrated. Interaction profiles of seven adhesion/growth‐regulatory galectins cover the range from intense signals with core 2 pentasaccharides and core 1 binding for galectins‐3 and ‐5 to a lack of binding for galectin‐1 and also the galectin‐related protein, which was included as a negative control. Remarkably, the two tandem‐repeat‐type galectins‐4 and ‐8 were distinguished by core 1 sialylation, as the two separated domains were. These results encourage further synthetic elaboration of the glycopeptide library and testing of the network of natural galectins and rationally engineered variants of the lectins. 相似文献
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Zhipeng Dong Qiyan Wang Ming Huo Nanxia Zhang Bingxia Li Hongmei Li Yisong Xu Meng Chen Prof. Hao Hong Prof. Yue Wang 《ChemistryOpen》2019,8(7):915-921
Dendritic cells (DCs) based cancer immunotherapy is largely dependent on adequate antigen delivery and efficient induction of DCs maturation to produce sufficient antigen presentation and ultimately lead to substantial activation of tumor-specific CD8+ T cells. Carbon nanotubes (CNTs) have attracted great attention in biomedicine because of their unique physicochemical properties. In order to effectively deliver tumor antigens to DCs and trigger a strong anti-tumor immune response, herein, a specific DCs target delivery system was assembled by using multi-walled carbon nanotubes modified with mannose which can specifically bind to the mannose receptor on DCs membrane. Ovalbumin (OVA) as a model antigen, could be adsorbed on the surface of mannose modified multi-walled carbon nanotubes (Man-MWCNTs) with a large drug loading content. This nanotube-antigen complex showed low cytotoxicity to DCs and was efficiently engulfed by DCs to induce DCs maturation and cytokine release in vitro, indicating that it could be a potent antigen-adjuvant nanovector of efficient antigen delivery for therapeutic purpose. 相似文献