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41.
Constructing a recombinant protein between a reporter enzyme and a detector protein to produce a homogeneous immunological reagent is advantageous over random chemical conjugation. However, the approach hardly recombines multiple enzymes in a difunctional fusion protein, which results in insufficient amplification of the enzymatic signal, thereby limiting its application in further enhancement of analytical signal. In this study, two site-specific biotinylated recombinant proteins, namely, divalent biotinylated alkaline phosphatase (AP) and monovalent biotinylated ZZ domain, were produced by employing the Avitag–BirA system. Through the high streptavidin (SA)–biotin interaction, the divalent biotinylated APs were clustered in the SA–biotin complex and then incorporated with the biotinylated ZZ. This incorporation results in the formation of a functional macromolecule that involves numerous APs, thereby enhancing the enzymatic signal, and in the production of several ZZ molecules for the interaction with immunoglobulin G (IgG) antibody. The advantage of this signal amplification strategy is demonstrated through ELISA, in which the analytical signal was substantially enhanced, with a 32-fold increase in the detection sensitivity compared with the ZZ–AP fusion protein approach. The proposed immunoassay without chemical modification can be an alternative strategy to enhance the analytical signals in various applications involving immunosensors and diagnostic chips, given that the label-free IgG antibody is suitable for the ZZ protein. 相似文献
42.
Dr. Radislav A. Potyrailo Anthony J. Murray Nandini Nagraj Andrew D. Pris Jeffrey M. Ashe Milos Todorovic 《Angewandte Chemie (International ed. in English)》2015,54(7):2174-2178
A single aptamer bioreceptor layer was formed using a common streptavidin–biotin immobilization strategy and employed for 100–365 bind/release cycles. Chemically induced aptamer unfolding and release of its bound target was accomplished using alkaline solutions with high salt concentrations or deionized (DI) water. The use of DI water scavenged from the ambient atmosphere represents a first step towards maintenance‐free biosensors that do not require the storage of liquid reagents. The aptamer binding affinity was determined by surface plasmon resonance and found to be almost constant over 100–365 bind/release cycles with a variation of less than 5 % relative standard deviation. This reversible operation of biosensors based on immobilized aptamers without storage of liquid reagents introduces a conceptually new perspective in biosensing. Such new biosensing capability will be important for distributed sensor networks, sensors in resource‐limited settings, and wearable sensor applications. 相似文献
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The paramagnetic contributions to the spin-lattice relaxation rates of biotin 13C nuclei, induced by the presence in the water/DMSO solution of the TEMPOL nitroxide, have been analysed in the interaction with avidin. The paramagnetic relaxation data, obtained at different temperatures, indicate that the average solvent/spin-label exposure of biotin carbons is consistent with the conformational features previously observed for the complex in the crystal. The analysis of the paramagnetic perturbation profiles, derived from 13C spin lattice relaxation measurements, seems to be highly informative of the sterical aspects of interaction processes of large biopolymers with their ligands. 相似文献
45.
The multilayer thin films composed of polymers, avidin, and concanavalin A (Con A) were prepared on the surface of a quartz slide by alternate deposition of anionic and cationic polymers and the proteins. The loading of avidin and Con A in the film was evaluated spectroscopically by using Texas Red tagged avidin and Con A. Avidin molecules assembled in the film caused its binding activity toward biotin and analogue 2-(4′-hydroxyphenylazo)benzoic acid to be retained. The polymer layers contributed to improving stability of the protein film.
46.
H. Morgan H. Fukushima D. M. Taylor 《Journal of polymer science. Part A, Polymer chemistry》1994,32(7):1331-1340
The synthesis of a range of aromatic bisbiotin ligands is described and their ability to polymerize the proteins avidin and streptavidin is investigated. The formation of protein polymers is confirmed by gel chromatography and transmission electron microscopy. It is shown that the degree of polymerization depends not only on the length of the ligand but also on the stereochemistry of the group linking the distal biotin moieties. © 1994 John Wiley & Sons, Inc. 相似文献
47.
E. A. Khatuntseva O. N. Yudina Yu. E. Tsvetkov A. A. Grachev R. N. Stepanenko R. Ya. Vlasenko V. L. Lvov N. E. Nifantiev 《Russian Chemical Bulletin》2006,55(11):2095-2102
Starting from the biotechnologically available trisaccharide sialyl-3′-lactose, representing the carbohydrate portion of the
tumor-associated ganglioside GM3, the corresponding 3-aminopropyl β-glycoside (1) and 3-(4-maleimidobutanoylamino)propyl glycoside were synthesized. The reaction of the latter with a thiolated derivative
of the Megathura crenulata hemocyanine (KLH) afforded a carbohydrate—protein conjugate, a tumor vaccine prototype containing about 330 trisaccharide
ligands attached to KLH. N-Stearoylation of ligand 1 gave the model neoglycolipid for comparative study of the activity of mono-and polyvalent immunogens and the natural ganglioside
GM3. A monovalent conjugate, in which ligand 1 is linked to biotin through an oligo(ethylene glycol) spacer and a polyvalent conjugate with a polyacrylamide carrier were
also prepared. These conjugates are meant as covering antigens to assess the specificity and efficiency of the immune response
in the ELISA assay.
Dedicated to Academician O. M. Nefedov on the occasion of his 75th birthday.
Published in Russian in Izvestiya Akademii Nauk. Seriya Khimicheskaya, No. 11, pp. 2016–2023, November, 2006. 相似文献
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49.
Dr. Mohammad Yaseen Ahmad Shuwen Liu Dr. Tirusew Tegafaw Abdullah Khamis Ali Al Saidi Dejun Zhao Ying Liu Sangyeol Lee Seungho Kim So Hyeon Yang Dong Wook Hwang Dr. Ahrum Baek Prof. Jae Chang Jung Prof. Sung-Wook Nam Prof. Kwon Seok Chae Prof. Yongmin Chang Prof. Gang Ho Lee 《欧洲无机化学杂志》2023,26(27):e202300430
Herein, biotin (Bio)-conjugated poly(acrylic acid) (PAA)-grafted ultrasmall gadolinium oxide nanoparticles (Bio-PAA-Gd2O3 NPs) were synthesized for enhanced tumor imaging using Bio as a tumor-targeting ligand. The average particle diameter of Gd2O3 NPs was 2.1 nm. The Bio-PAA-Gd2O3 NPs exhibited excellent colloidal stability (i. e., no precipitation) and a high longitudinal water proton spin relaxivity (r1) of 23.8 s−1 mM−1 (r2/r1=1.6 and r2=transverse water proton spin relaxivity), which was ∼6 times higher than those of commercial Gd-chelated magnetic resonance imaging (MRI) contrast agents. Cytotoxicity tests using two cell lines showed that the Bio-PAA-Gd2O3 NPs were almost non-toxic up to the measured concentration of 500 μM Gd. The enhanced tumor imaging of the Bio-PAA-Gd2O3 NPs was demonstrated through their higher positive contrasts and longer contrast retention at the tumor after intravenous injection in T1 MR images, compared with those of the control PAA-Gd2O3 NPs. 相似文献
50.