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The chemical diversity and structural flexibility of lead halide perovskites (LHPs) offer tremendous opportunities to tune their optical properties through internal molecular engineering and external stimuli. Herein, we report the wide-range and ultrapure photoluminescence emissions in a family of homologous 2D LHPs, [MeOPEA]2 PbBr4-4x I4x (MeOPEA=4-methoxyphenethylammonium; x=0, 0.2, 0.425, 0.575, 1) enabled through internal chemical pressure and external hydrostatic pressure. The chemical pressure, induced by the C-Hâ â â π interactions and halogen doping/substitution strengthens the structural rigidity to give sustained narrow emissions, and regulates the emission energy, respectively. Further manipulation of physical pressure leads to wide-range emission tuning from 412 to 647â nm in a continuous and reversible manner. This work could open up new pathways for developing 2D LHP emitters with ultra-wide color gamut and high color purity which are highly useful for pressure sensing.
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Nanobodies show a great potential in biomedical and biotechnology applications. Bacterial expression is the most widely used expression system for nanobody production. However, the yield of nanobodies is relatively low compared to that of eukaryotic systems. In this study, the repetitive amino acid sequence motifs (GAGAGS) found in silk fibroin protein (SFP) were developed as a novel fusion tag (SF-tag) to enhance the expression of nanobodies in Escherichia coli. SF-tags of 1 to 5 hexapeptide units were fused to the C-terminus of 4G8, a nanobody against human epididymis protein 4 (HE4). The protein yield of 4G8 variants was increased by the extension of hexapeptide units and achieved a 2.5 ~ 7.1-fold increase compared with that of untagged 4G8 (protein yield of 4G8-5C = 0.307 mg/g vs that of untagged 4G8 = 0.043 mg/g). Moreover, the fusion of SF-tags not only had no significant effect on the affinity of 4G8, but also showed a slight increase in the thermal stability of SF-tag-fused 4G8 variants. The fusion of SF-tags increased the transcription of 4G8 by 2.3 ~ 7.0-fold, indicating SF-tags enhanced the protein expression at the transcriptional level. To verify the applicability of the SF-tags for other nanobody expression, we further investigated the protein expression of two other anti-HE4 nanobodies 1G8 and 3A3 upon fusion with the SF-tags. Results indicated that the SF-tags enhanced the protein expression up to 5.2-fold and 5.7-fold for 1G8 and 3A3, respectively. For the first time, this study reported a novel and versatile fusion tag system based on the SFP for improving nanobody expression in Escherichia coli, which may enhance its potential for wider applications.Key points⢠A silk fibroin protein-based fusion tag (SF-tag) was developed to enhance the expression of nanobodies in Escherichia coli.⢠The SF-tag enhanced the nanobody expression at the transcriptional level.⢠The fusion of SF-tag had no significant effect on the affinity of nanobodies and could slightly increase the thermal stability of nanobodies.
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Fibroínas , Anticorpos de Domínio Único , Biotecnologia , Escherichia coli/genética , Escherichia coli/metabolismo , Fibroínas/genética , Humanos , Processamento de Proteína Pós-Traducional , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Anticorpos de Domínio Único/químicaRESUMO
The high-pressure and high-temperature behaviors of manganotantalite Mn(Ta,Nb)2O6 have been investigated by single-crystal X-ray diffraction and Raman spectroscopy combined with diamond anvil cell technique, as well as first-principle calculations. A pressure-induced reversible phase transition of manganotantalite occurs at 9.5 GPa and room temperature, accompanied by a large volume collapse (â¼7.0%) and drastic color change from brownish-yellow to red. The space groups of low-pressure (LP) and high-pressure (HP) phases are the same (Pbcn), but the coordination numbers of Mn increase from six to eight and Ta increase from six to seven, respectively. The band gap becomes narrow from 2.37 to 1.59 eV. We determined the P-T phase diagram of manganotantalite with a positive Clapeyron slope of dP/dT = 0.0073 GPa/K. The P-V data were fitted to a second-order Birch-Murnaghan equation of state with B0 = 149(4) GPa for the LP phase and B0 = 188(3) GPa for the HP phase. The isothermal Grüneisen parameters were determined to be 0.23â¼2.03 of the LP phase and 0.59â¼0.86 of the HP phase for Raman modes. High-pressure behaviors of Mn(Ta,Nb)2O6 indicate that this kind of material is a potential effective pressure sensor to monitor pressure change or warn pressure abnormality.
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Carcinoembryonic antigen-related cell adhesion molecule 5 (CEACAM-5) assays are employed in routine clinical settings to diagnose tumor. We selected two nanobodies with high-affinity to CEACAM-5, termed Nb11C12 and Nb2D5, using phage-display technology. The Nb2D5 fused with calf intestinal alkaline phosphatase (CAP), human placental alkaline phosphatase (HAP), or Pyrococcus abyssi alkaline phosphatase (PAP) were expressed in human embryonic kidney (HEK293) cells. The enzymatic activity of Nb2D5-HAP fusion protein was the best and remained stable at 60 °C for 7 days. The affinity of Nb2D5-HAP fusion protein to CEACAM-5 reached 42 pM. A chemiluminescent enzyme immunoassay (CLEIA) based on Nb2D5-HAP fusion protein was established for quantitative CEACAM-5 assay in clinical settings. The CLEIA exhibited a wide linear range of 0.31-640 ng/mL toward CEACAM-5, with a limit of detection (LOD) of 0.85 ng/mL. No cross-reactivity occurred with CEACAM-1, CEACAM-3, CEACAM-6, or CEACAM-8, and no interference was observed with rheumatoid factors. The CLEIA based on Nb2D5-HAP fusion protein was stable for 8 weeks at 37 °C and 50% relative humidity. The CLEIA developed from Nb2D5-HAP fusion protein had much better stability and linearity with similar reproducibility compared with the enzyme-linked immunosorbent assay developed from conventional monoclonal antibodies, which have been widely used in clinics over the past several decades. Graphical abstract.
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Fosfatase Alcalina/metabolismo , Antígeno Carcinoembrionário/metabolismo , Técnicas Imunoenzimáticas/métodos , Proteínas Recombinantes de Fusão/metabolismo , Anticorpos de Domínio Único , Antígeno Carcinoembrionário/imunologia , Proteínas Ligadas por GPI/imunologia , Células HEK293 , Humanos , Limite de Detecção , Luminescência , Reprodutibilidade dos TestesRESUMO
Nerve growth factor (NGF) is an essential trophic factor for the growth and survival of neurons in the central and peripheral nervous systems. For many years, mouse NGF (mNGF) has been used to treat various neuronal and non-neuronal disorders. However, the biological activity of human NGF (hNGF) is significantly higher than that of mNGF in human cells. Using the CRISPR/Cas9 system, we constructed the transgenic mice expressing hNGF specifically in their submandibular glands. As demonstrated by fluorescence immunohistochemical staining, these mice produced hNGF successfully, with 0.8 mg produced per gram of submandibular glands. hNGF with 99% purity was successfully extracted by two-step ion-exchange chromatography and one-step size-exclusion chromatography from the submandibular glands of these transgenic mice. Further, the purified hNGF was verified by LC-MS/MS. We analyzed the NH2-terminus of hNGF using both Edman degradation and LC-MS/MS-based methods. Both results showed that the obtained hNGF lost the NH2-terminal octapeptide (SSSHPIFH). Moreover, the produced hNGF demonstrated a strong promotion in the proliferation of TF1 cells.
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Edição de Genes/métodos , Fator de Crescimento Neural/isolamento & purificação , Fator de Crescimento Neural/metabolismo , Glândula Submandibular/metabolismo , Animais , Linhagem Celular , Proliferação de Células , Cromatografia em Gel , Cromatografia por Troca Iônica , Humanos , Camundongos , Camundongos Transgênicos , Fator de Crescimento Neural/química , Fator de Crescimento Neural/genética , Domínios Proteicos , Engenharia de ProteínasRESUMO
Nanobodies offer multiple advantages over conventional antibodies in terms of size, stability, solubility, immunogenicity, and production costs, with improved tumor uptake and blood clearance. Additionally, the recombinant expression of nanobodies is robust in various expression systems, such as Escherichia coli, Saccharomyces cerevisiae and Pichia pastoris. P. pastoris is the most widely used microorganism for nanobody production, but all or almost all expression vectors developed for this system are based on the regulated promoter of the alcohol oxidase 1 gene (AOX1) that requires methanol for full induction. In this study, a constitutive anti-CEACAM5 nanobody expression system was constructed under the control of a glyceraldehyde-3-phosphate dehydrogenase promoter (GAP) promoter. The effects of different carbon sources and pH on nanobody expression were evaluated in shaking flask cultures. After 96â¯h of constitutive expression in shaking flask, a yield of 51.71â¯mg/L was obtained. In addition, this constitutive expression system produced nanobodies at equivalent yield and affinity to that produced by methanol-induced expression. The results of this study indicated that the use of a constitutive expression system is a promising alternative for the production of nanobodies applied for cancer diagnosis and therapy.
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Vetores Genéticos/genética , Pichia/genética , Anticorpos de Domínio Único/genética , Animais , Técnicas de Cultura Celular por Lotes , Camelídeos Americanos , Antígeno Carcinoembrionário/imunologia , Proteínas Ligadas por GPI/imunologia , Expressão Gênica , Gliceraldeído-3-Fosfato Desidrogenases/genética , Humanos , Regiões Promotoras Genéticas , Proteínas Recombinantes/genética , Anticorpos de Domínio Único/imunologiaRESUMO
Broadband emission in hybrid lead halide perovskites (LHPs) has gained significant attention due to its potential applications in optoelectronic devices. The origin of this broadband emission is primarily attributed to the interactions between electrons and phonons. Most investigations have focused on the impact of structural characteristics of LHPs on broadband emission, while neglecting the role of electronic mobility. In this work, the study investigates the electronic origins of broadband emission in a family of 2D LHPs. Through spectroscopic experiments and density functional theory calculations, the study unveils that the electronic states of the organic ligands with conjugate effect in LHPs can extend to the band edges. These band-edge carriers are no longer localized only within the inorganic layers, leading to electronic coupling with molecular states in the barrier and giving rise to additional interactions with phonon modes, thereby resulting in broadband emission. The high-pressure photoluminescence measurements and theoretical calculations reveal that hydrostatic pressure can induce the reconfiguration of band-edge states of charge carriers, leading to different types of band alignment and achieving macroscopic control of carrier dynamics. The findings can provide valuable guidance for targeted synthesis of LHPs with broadband emission and corresponding design of state-of-the-art optoelectronic devices.
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BACKGROUND: Carcinoembryonic antigen-related cell adhesion molecule 5 (CEACAM-5) is a well-characterized biomarker for the clinical diagnosis of various cancers. Nanobodies, considered the smallest antibody fragments with intact antigen-binding capacity, have gained significant attention in disease diagnosis and therapy. Due to their peculiar properties, nanobodies have become promising alternative diagnostic reagents in immunoassay. However, nanobodies-based immunoassay is still hindered by small molecular size and low antigen capture efficacy. Therefore, there is a pressing need to develop novel nanobody-based immunoassays with superior performance. RESULTS: A novel pentameric nanobodies-based immunoassay (PNIA) was developed with enhanced sensitivity and specificity for CEACAM-5 detection. The binding epitopes of three anti-CEACAM-5 nanobodies (Nb1, Nb2 and Nb3) were analyzed. To enhance the capture and detection efficacy of CEACAM-5 in the immunoassay, we engineered bispecific nanobodies (Nb1-Nb2-rFc) as the capture antibody, and developed the FITC-labeled pentameric nanobodies (Nb3-VT1B) as the detection antibody. The binding affinities of Nb1-Nb2-rFc (1.746 × 10-10) and Nb3-VT1B (1.279 × 10-11) were significantly higher than those of unmodified nanobodies (Nb1-rFc, 4.063 × 10-9; Nb2-rFc, 2.136 × 10-8; Nb3, 3.357 × 10-9). The PNIA showed a linear range of 0.625-160 ng mL-1 with a correlation coefficient R2 of 0.9985, and a limit of detection of 0.52 ng mL-1, which was 24-fold lower than the immunoassay using monomeric nanobody. The PNIA was validated with the spiked human serum. The average recoveries ranged from 91.8% to 102% and the coefficients of variation ranged from 0.026% to 0.082%. SIGNIFICANCE AND NOVELTY: The advantages of nanobodies offer a promising alternative to conventional antibodies in disease diagnosis. The novel PNIA demonstrated superior sensitivity and high specificity for the detection of CEACAM-5 antigen. This bispecific or multivalent nanobody design will provide some new insights into the design of immunoassays for clinical diagnosis.
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Anticorpos de Domínio Único , Humanos , Antígeno Carcinoembrionário , Imunoensaio , Anticorpos , Sensibilidade e Especificidade , Ensaio de Imunoadsorção EnzimáticaRESUMO
The Raman spectra and thermal expansion of MgP2O6 metaphosphate were investigated at various temperatures up to 1073 K at ambient pressure. No temperature-induced phase transition was observed in this study. The effect of temperature on the Raman active vibrations and unit cell parameters was determined. All the observed Raman active bands of MgP2O6 showed linear temperature dependences in the range of -2.61 × 10-2 â¼ -0.49 × 10-2 cm-1 K-1. The thermal expansion coefficient of MgP2O6 was estimated to be 3.21(2) × 10-5 K-1. An axial anisotropic thermal expansion exists and the c-axis shows the smallest thermal expansion. The isobaric mode Grüneisen parameters of MgP2O6 were calculated. The obtained results were compared with other compounds in the MgO-P2O5 system.
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This work aimed to establish a simple and feasible method to obtain silk fibroin nanoparticles (SFNPs) with uniform particles size, and then modify the SFNPs with nanobody (Nb) 11C12 targeting the proximal membrane end of carcinoembryonic antigen on the surface of colorectal cancer (CRC) cells. The regenerated silk fibroin (SF) was isolated using ultrafiltration tubes with a 50 kDa molecular weight cut-off, and the retention fraction (named as SF > 50 kDa) was further self-assembled into SFNPs by ethanol induction. Scanning electron microscope (SEM) and high-resolution transmission electron microscop showed that the SFNPs with uniform particles size were formed. Due to electrostatic adsorption and pH responsiveness, SFNPs have been proved to effectively load and release the anticancer drug doxorubicin hydrochloride (DOX) (DOX@SFNPs). Further, targeting molecule Nb 11C12 was used to modify these nanoparticles, constituting the targeted outer layer of the drug delivery system (DOX@SFNPs-11C12), achieving precise localization to cancer cells. The release amount of DOX observed fromin vitrodrug release profiles increased as follows: pH 7.4 < pH 6.8 < pH 5.4, demonstrating that the DOX release could be accelerated in a weakly acidic environment.In vitrocytotoxicity experiments displayed that SFNPs-11C12 nanoparticles exhibited good safety and biocompatibility. Drug-loaded nanoparticles, DOX@SFNPs-11C12, led to higher LoVo cells apoptosis compared to DOX@SFNPs. Fluorescence spectrophotometer characterization and confocal laser scanning microscopy further showed that the internalization of DOX was highest in the DOX@SFNPs-11C12, certifying that the introduced targeting molecule enhanced the uptake of drug delivery system by LoVo cells. This study provides a simple and operational approach to developing an optimized SFNPs drug delivery system modified by targeting Nb, which can be a good candidate for CRC therapy.
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Antineoplásicos , Neoplasias Colorretais , Fibroínas , Nanopartículas , Humanos , Fibroínas/química , Antígeno Carcinoembrionário , Nanopartículas/química , Doxorrubicina/farmacologia , Sistemas de Liberação de Medicamentos , Concentração de Íons de HidrogênioRESUMO
microRNAs (miRNAs or miRs) have been reported to regulate the pathology of intracerebral hemorrhage (ICH). Therefore, the present study aimed to investigate the function of miR-30e-5p in rats with ICH with specific focus on Toll-like receptor (TLR)4. In the present study, collagenase type IV was used for the establishment of the ICH model in rats, prior to which the rats were injected with miR-30e-5p mimic or miR-30e-5p mimic + pcDNA3.1-TLR4 plasmid. The expression levels of miR-30e-5p and TLR4 were then measured using reverse transcription-quantitative PCR and western blotting. The potential interaction between miR-30e-5p and TLR4 was tested using the MicroRNA Target Prediction Database and dual-luciferase reporter and RNA immunoprecipitation assay. In addition, the concentration of TNF-α, IL-6 and IL-1ß was measured using ELISA. The protein expression levels of TLR4/myeloid differentiation factor 88 (MyD88)/TIR-domain-containing adapter-inducing interferon-ß (TRIF) signaling-associated molecules were measured by western blotting. Following induction of ICH, miR-30e-5p expression was downregulated, while TLR4 expression was upregulated. By contrast, injection with miR-30e-5p mimic rescued neuronal function while suppressing neuronal inflammation in rats following ICH; these effects were reversed by co-overexpression of TLR4. Furthermore, overexpression of miR-30e-5p inactivated TLR4/MyD88/TRIF signaling in rats with ICH; this was also reversed by overexpression of TLR4. Taken together, these results suggested that overexpression of miR-30e-5p exerted a protective role against neuronal deficit and inflammation caused by ICH in rats by targeting TLR4 and inactivating TLR4/MyD88/TRIF signaling.
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Controllable protein attachment onto solid interfaces is essential for the functionality of proteins with broad applications. Silica-binding peptides (SBPs) have emerged as an important tool enabling convenient binding of proteins onto a silica surface. Surprisingly, we found that removal of polyhistidines, a common tag for protein purification, dramatically decrease the binding affinity of a SBP-tagged nanobody onto a silica surface. We hypothesized that polyhistidines and SBPs can be combined to enhance affinity. Through a series of purposely designed SBPs, we identified that the relative orientation of amino acids is a key factor affecting the surface binding strength. One re-engineered SBP, SBP4, exhibits a 4000-fold improvement compared to the original sequence. Guided by physical insights, the work provides a simple strategy that can dramatically improve affinity between a SBP and a silica surface, promising a new way for controllable immobilization of proteins, as demonstrated using nanobodies.
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Histidina/química , Proteínas/química , Dióxido de Silício/química , Sequência de Aminoácidos , Modelos Moleculares , Ligação Proteica , Conformação Proteica , Propriedades de SuperfícieRESUMO
Human epididymis protein 4 (HE4) is a glycoprotein secreted by epithelial ovarian cancer (EOC) cells and is a novel and specific biomarker for diagnosing and prognosing EOC. Previous studies have shown that overexpression of HE4 is correlated with EOC tumorigenesis and chemoresistance. However, less has been reported regarding the direct effect of the secreted HE4 protein as an autocrine factor in EOC cells. Here, we investigated the molecular mechanism of the secretory form of HE4 on the growth of EOC cells by applying nanobodies with a targeted interaction of free HE4. Three anti-HE4 nanobodies were selected from an immune library by phage display. HE4 secreted by serum-free cultured OVCAR3 cells increased and was effectively neutralized by anti-HE4 nanobodies, which inhibited cell viability. Treatment with the anti-HE4 nanobody 1G8 decreased Bcl-2 expression and increased BAX, cleaved PARP, and p53 levels, resulting in apoptosis of OVCAR3 cells. Moreover, 1G8 significantly improved the cisplatin response of OVCAR3 cells. Our data suggest that secretory HE4 played a novel pro-survival autocrine role and was a target of the anti-HE4 nanobody to improve the therapeutic effects of cisplatin-based chemotherapy.
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Neoplasias Epiteliais e Glandulares , Neoplasias Ovarianas , Anticorpos de Domínio Único , Apoptose , Biomarcadores Tumorais/uso terapêutico , Carcinoma Epitelial do Ovário/tratamento farmacológico , Linhagem Celular Tumoral , Cisplatino/farmacologia , Cisplatino/uso terapêutico , Feminino , Humanos , Neoplasias Epiteliais e Glandulares/tratamento farmacológico , Neoplasias Ovarianas/metabolismo , Anticorpos de Domínio Único/farmacologiaRESUMO
Uniaxial and notched tension samples are utilized to investigate the damage and failure of titanium alloy Ti6Al4V. The strain fields on the samples are obtained by the digital image correlation (DIC) method. Strain localization occurs before fracturing in all samples, and the width and size of the localized zone are characterized. Slant fractures are observed in uniaxial and notched tension specimen, which indicate that the initiation and propagation of cracks in thin sheet specimens are highly affected by the shear stress. Numerical simulations were performed for identification of hybrid hardening laws, and the results were compared with the experiments. The influence of the stress triaxiality on damage mechanism of Ti6Al4V was analyzed by observation of the specimen fracture surfaces using SEM. The results show that a higher stress triaxiality facilitates the formation and growth of micro-voids, which leads to a decrement of strain at failure.
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Recently, injectable hydrogels have shown great potential in cell therapy and drug delivery. They can easily fill in any irregular-shaped defects and remain in desired positions after implantation using minimally invasive strategies. Here, we developed hydrogels prepared from tilapia skin collagen and chitosan (HCC). The residual mass rate of HCC was affected by the pH at the time of preparation, which was 29.1 % at pH 7 in 36â¯h. By comparison, the residual mass ratios of HCC at pH values of 6 and 5 were only approximately 8.4 % and 0, respectively. In addition, the stability of HCC was also affected by the concentration of these two components. HCC10 catalyzed by 10â¯mgâ¯mL-1 tilapia skin collagen and 10â¯mgâ¯mL-1 chitosan was more stable than HCC5 catalyzed by 5â¯mgâ¯mL-1 tilapia skin collagen and 10â¯mgâ¯mL-1 chitosan; therefore, we studied that ability of HCC10 to deliver two model nanobodies: 2D5 and KPU. As the concentration of nanobodies increased, the cumulative release rate of 2D5 decreased, and the release rate of KPU increased. Meanwhile, the cumulative release rate of 2D5 was the highest (68.3 %) at pH 5.5, followed by pH 6.8 (56.4 %) and 7.4 (28.4 %). However, the cumulative release rates of KPU were similar at pH 5.5 (45.1 %), 6.8 (46.5 %), and 7.4 (44.9 %). HCC is biodegradable, and can facilitate the release nanobodies; thus, HCC could be developed into an intelligent responsive tumor treatment matrix for use in cancer therapy.
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Quitosana , Tilápia , Animais , Colágeno , Hidrogéis , Concentração de Íons de HidrogênioRESUMO
Progressive depositions of cerebral amyloid are primary neuropathologic features of Alzheimer's disease (AD). The amyloid is composed of a 39-42 amino acid peptide called the amyloid beta-protein (Abeta). Repeated investigation suggests that the conformational transition of Abeta from alpha-helix or random coil to beta-sheet structure plays a key role in the inappropriate accumulation of cerebral amyloid plaques. In this manuscript, we describe a fluorescence-based immunoassay technology to investigate the conformation and topology of Abeta peptides interacting with peptide-tethered planar lipid bilayers. Dual monoclonal antibodies (mAbs) labelled with fluorophores were employed to recognise a linear N- and a beta-sheet C-terminus of Abeta peptides on the model membrane, respectively. Kinetics of antibody-Abeta binding were determined by surface plasmon field-enhanced fluorescence spectroscopy (SPFS). The conformational transition of Abeta by melatonin, a defined beta-sheet breaker, was probed using paired monoclonal antibodies. The Abeta interaction with the membrane was evaluated by carefully analyzing the change in kinetic/affinity parameters in the presence or absence of melatonin. These results show that SPFS can be used to examine conformational transition of Abeta on an artificial membrane, providing a novel and versatile platform for conveniently monitoring protein-membrane interaction and screening for new beta-sheet breakers.
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Peptídeos beta-Amiloides/química , Membranas Artificiais , Absorção , Bicamadas Lipídicas/química , Modelos Biológicos , Espectrometria de Fluorescência , Ressonância de Plasmônio de SuperfícieRESUMO
Positively charged and pH self-buffering quantum dots (Tren-QDs) were achieved by surface functionalization with tris(2-aminoethyl)amine (Tren) derivatives, which are attached to the inorganic cores of QDs through bidentate chelating of dithiocarbamates. The Tren-QDs exhibit pH buffering capability by absorbing or releasing protons due to the surface polyamine groups as the surrounding pH fluctuates. Such self-buffering capability stabilizes the photoluminescence of the Tren-QDs against acid. The Tren-QDs bear positive charges through protonation of the surface polyamine groups under physiological conditions and the surface positive charges improve their cellular uptake efficiency by charge mediation, which has been demonstrated by BV-2 microglia cells. The photoluminescence of Tren-QDs shows a selective Stern-Volmer response to copper ions and this property has been preliminarily evaluated for investigating the BV-2 cell membrane structure by monitoring the photoluminescence of intracellular Tren-QDs.
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Permeabilidade da Membrana Celular/fisiologia , Poliaminas/química , Pontos Quânticos , Ácido 3-Mercaptopropiônico/química , Ácido 3-Mercaptopropiônico/farmacocinética , Animais , Soluções Tampão , Linhagem Celular , Membrana Celular/metabolismo , Cobre/química , Etilenodiaminas/química , Etilenodiaminas/farmacocinética , Concentração de Íons de Hidrogênio , Luminescência , Camundongos , Microglia/metabolismo , Microscopia Eletrônica de Transmissão , Compostos Organofosforados/química , Compostos Organofosforados/farmacocinética , Poliaminas/farmacocinética , Espectrofotometria Ultravioleta , Propriedades de SuperfícieRESUMO
This paper investigates the fatigue damage and cracking behavior of aluminum alloy 2024-T4 with different levels of prior corrosion. Damage evolution, crack initiation and propagation were experimentally analyzed by digital image correlation, scanning electron microscopy and damage curves. Prior corrosion is shown to cause accelerated damage accumulation, inducing premature fatigue crack initiation, and affecting crack nucleation location, crack orientation and fracture path. For the pre-corrosion condition, although multiple cracks were observed, only one corrosion-initiated primary crack dominates the failure process, in contrast to the plain fatigue cases, where multiple cracks propagated simultaneously leading to final coalescence and fracture. Based on the experimental observations, a mixed-mode fracture model is proposed and shown to successfully predict fatigue crack growth and failure from the single dominant localized corrosion region.
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Nanobodies represent the next-generation antibody-derived biologics with significant advances over conventional antibodies. Several rapid and robust techniques for isolating highly specific nanobodies have been developed. Antigen specific nanobodies are selected from constructed nanobody libraries, which can be classified into 3 main types: immune library, naïve library, and semisynthetic/synthetic library. The immune library is the most widely used strategy for nanobody screening. Target specific nanobodies are highly enriched in immune libraries than in non-immune libraries; however, it is largely limited by the natural antigenicity of antigens. The naïve library is thus developed. Despite the lack of somatic maturation, protein engineering can be employed to significantly increase the affinities of selected binders. However, a substantial amount of blood samples collected from a large number of individual animals is a prerequisite to ensure the diversity of the naïve library. With this issue considered, the semisynthetic/synthetic library may be a promising path toward obtaining a limitless source of nanobodies against a variety of antigens without the need of animals. In this review, we summarize the state-of-the-art screening technologies with different libraries. The approaches presented here can further boost the diverse applications of nanobodies in biomedicine and biotechnology.
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Anticorpos de Domínio Único , Animais , Humanos , Biblioteca de PeptídeosRESUMO
OBJECTIVE: To investigate the anti-lipid peroxidation of the water extract from Cornu Cervi Pantotrichum (WECCP) in vitro. METHODS: The lipid peroxidation (malondialdehyde formation) in microsomes of rat brain, liver and kidney induced by NADPH-Vit C and cyctein-Fe2+, and the superoxide anion radical production (reduced cytochrome c formation) in xanthine-xanthine oxidase system in vitro were evaluated. RESULTS: The lipid peroxidation (MDA formation) in microsomes of rat brain, liver and kidney induced by NADPH-Vit C and cyctein-Fe2+, O2 production(reduced cytochrome c formation) in xanthine-xanthine oxidase system were significantly inhibited by WECCP. CONCLUSION: WECCP exhibits an antioxidant activity.