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https://www.readbyqxmd.com/read/29457451/boosted-electrochemical-immunosensing-gm-crops-markers-using-nanobody-and-mesoporous-carbon
#1
Mingming Zhang, Guanghui Li, Qing Zhou, Deng Pan, Min Zhu, Runyu Xiao, Yuanjian Zhang, Guoqiu Wu, Yakun Wan, Yanfei Shen
The problems of environmental security and the potential risks of human health caused by transgenic crops have aroused numerous attention. Recent studies reveal 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) from Agrobacterium sp. strain CP4 protein (CP4-EPSPS), which shows very high resistance to herbicide glyphosate, is a typical biomarker of genetically modified (GM) crops. For this reason, it is highly anticipated to devise a sensitive and convenient strategy to detect CP4-EPSPS protein in crops. Herein, we reported a simple electrochemical immunosensor by coupling nanobody, ordered mesoporous carbon (OMC), and thionine (Th)...
February 19, 2018: ACS Sensors
https://www.readbyqxmd.com/read/29444803/open-source-recombinant-monoclonal-secondary-nanobodies
#2
Helge Ewers
Secondary antibodies are everyday reagents in biomedical research that are generated in animals. In this issue, Pleiner et al. (2018. J. Cell Biol. https://doi.org/10.1083/jcb.201709115) describe several single domain antibody fragments against antibodies from mouse and rabbit, so-called nanobodies that are easily produced recombinantly, and characterize their use in Western blotting, enzyme-linked immunosorbent assay, and immunofluorescence assays.
February 14, 2018: Journal of Cell Biology
https://www.readbyqxmd.com/read/29443069/the-use-of-a-%C3%AE-lactamase-based-conductimetric-biosensor-assay-to-detect-biomolecular-interactions
#3
Marylène Vandevenne, Mathieu Dondelinger, Sami Yunus, Astrid Freischels, Régine Freischels, Oscar Crasson, Noureddine Rhazi, Pierre Bogaerts, Moreno Galleni, Patrice Filée
Biosensors are becoming increasingly important and implemented in various fields such as pathogen detection, molecular diagnosis, environmental monitoring, and food safety control. In this context, we used β-lactamases as efficient reporter enzymes in several protein-protein interaction studies. Furthermore, their ability to accept insertions of peptides or structured proteins/domains strongly encourages the use of these enzymes to generate chimeric proteins. In a recent study, we inserted a single-domain antibody fragment into the Bacillus licheniformis BlaP β-lactamase...
February 1, 2018: Journal of Visualized Experiments: JoVE
https://www.readbyqxmd.com/read/29440677/nanobody-triggered-lockdown-of-vsrs-reveals-ligand-reloading-in-the-golgi
#4
Simone Früholz, Florian Fäßler, Üner Kolukisaoglu, Peter Pimpl
Protein degradation in lytic compartments is crucial for eukaryotic cells. At the heart of this process, vacuolar sorting receptors (VSRs) bind soluble hydrolases in the secretory pathway and release them into the vacuolar route. Sorting efficiency is suggested to result from receptor recycling. However, how and to where plant VSRs recycle remains controversial. Here we present a nanobody-epitope interaction-based protein labeling and tracking approach to dissect their anterograde and retrograde transport routes in vivo...
February 13, 2018: Nature Communications
https://www.readbyqxmd.com/read/29440485/reporter-nanobody-fusions-ranbodies-as-versatile-small-sensitive-immunohistochemical-reagents
#5
Masahito Yamagata, Joshua R Sanes
Sensitive and specific antibodies are essential for detecting molecules in cells and tissues. However, currently used polyclonal and monoclonal antibodies are often less specific than desired, difficult to produce, and available in limited quantities. A promising recent approach to circumvent these limitations is to employ chemically defined antigen-combining domains called "nanobodies," derived from single-chain camelid antibodies. Here, we used nanobodies to prepare sensitive unimolecular detection reagents by genetically fusing cDNAs encoding nanobodies to enzymatic or antigenic reporters...
February 13, 2018: Proceedings of the National Academy of Sciences of the United States of America
https://www.readbyqxmd.com/read/29434474/suppression-of-breast-cancer-cell-proliferation-by-selective-single-domain-antibody-for-intracellular-stat3
#6
Sunanda Singh, Genoveva Murillo, Dong Chen, Ashutosh S Parihar, Rajendra G Mehta
Background: The serendipitous discovery of heavy-chain antibodies devoid of light chains in camelids and the subsequent development of VHHs (variable region of camelid heavy chain) have provided a very important tool for research and possibly for therapeutics. In this study, we synthesized single-domain 15-kDa antibody SBT-100 (anti-STAT3 B VHH13) against human STAT3 (signal transducer and activator of transcription) that binds selectively to STAT3 and suppresses the function of phosphorylated STAT3 (p-STAT3)...
2018: Breast Cancer: Basic and Clinical Research
https://www.readbyqxmd.com/read/29434346/yeast-surface-display-platform-for-rapid-discovery-of-conformationally-selective-nanobodies
#7
Conor McMahon, Alexander S Baier, Roberta Pascolutti, Marcin Wegrecki, Sanduo Zheng, Janice X Ong, Sarah C Erlandson, Daniel Hilger, Søren G F Rasmussen, Aaron M Ring, Aashish Manglik, Andrew C Kruse
Camelid single-domain antibody fragments ('nanobodies') provide the remarkable specificity of antibodies within a single 15-kDa immunoglobulin V HH domain. This unique feature has enabled applications ranging from use as biochemical tools to therapeutic agents. Nanobodies have emerged as especially useful tools in protein structural biology, facilitating studies of conformationally dynamic proteins such as G-protein-coupled receptors (GPCRs). Nearly all nanobodies available to date have been obtained by animal immunization, a bottleneck restricting many applications of this technology...
February 12, 2018: Nature Structural & Molecular Biology
https://www.readbyqxmd.com/read/29415506/vhh-antibodies-reagents-for-mycotoxin-detection-in-food-products
#8
REVIEW
Jia Wang, Hina Mukhtar, Lan Ma, Qian Pang, Xiaohong Wang
Mycotoxins are the toxic secondary metabolites produced by fungi and they are a worldwide public health concern. A VHH antibody (or nanobody) is the smallest antigen binding entity and is produced by heavy chain only antibodies. Compared with conventional antibodies, VHH antibodies overcome many pitfalls typically encountered in clinical therapeutics and immunodiagnostics. Likewise, VHH antibodies are particularly useful for monitoring mycotoxins in food and feedstuffs, as they are easily genetic engineered and have superior stability...
February 6, 2018: Sensors
https://www.readbyqxmd.com/read/29415042/comparing-autotransporter-%C3%AE-domain-configurations-for-their-capacity-to-secrete-heterologous-proteins-to-the-cell-surface
#9
Wouter S P Jong, Maaike Schillemans, Corinne M Ten Hagen-Jongman, Joen Luirink, Peter van Ulsen
Monomeric autotransporters have been extensively used for export of recombinant proteins to the cell surface of Gram-negative bacteria. A bottleneck in the biosynthesis of such constructs is the passage of the outer membrane, which is facilitated by the β-domain at the C terminus of an autotransporter in conjunction with the Bam complex in the outer membrane. We have evaluated eight β-domain constructs for their capacity to secrete fused proteins to the cell surface. These constructs derive from the monomeric autotransporters Hbp, IgA protease, Ag43 and EstA and the trimeric autotransporter Hia, which all were selected because they have been previously used for secretion of recombinant proteins...
2018: PloS One
https://www.readbyqxmd.com/read/29401625/novel-half-life-extended-anti-mif-nanobodies-protect-against-endotoxic-shock
#10
Amanda Sparkes, Patrick De Baetselier, Lea Brys, Inês Cabrito, Yann G-J Sterckx, Steve Schoonooghe, Serge Muyldermans, Geert Raes, Richard Bucala, Peter Vanlandschoot, Jo A Van Ginderachter, Benoît Stijlemans
Sepsis-leading to septic shock-is the leading cause of death in intensive care units. The systemic inflammatory response to infection, which is initiated by activated myeloid cells, plays a key role in the lethal outcome. Macrophage migration inhibitory factor (MIF) is an upstream immunoregulatory mediator, released by myeloid cells, that underlies a common genetic susceptibility to different infections and septic shock. Accordingly, strategies that are aimed at inhibiting the action of MIF have therapeutic potential...
January 25, 2018: FASEB Journal: Official Publication of the Federation of American Societies for Experimental Biology
https://www.readbyqxmd.com/read/29385713/nanobody-based-dual-specific-cars
#11
Stijn De Munter, Joline Ingels, Glenn Goetgeluk, Sarah Bonte, Melissa Pille, Karin Weening, Tessa Kerre, Hinrich Abken, Bart Vandekerckhove
Recent clinical trials have shown that adoptive chimeric antigen receptor (CAR) T cell therapy is a very potent and possibly curative option in the treatment of B cell leukemias and lymphomas. However, targeting a single antigen may not be sufficient, and relapse due to the emergence of antigen negative leukemic cells may occur. A potential strategy to counter the outgrowth of antigen escape variants is to broaden the specificity of the CAR by incorporation of multiple antigen recognition domains in tandem...
January 30, 2018: International Journal of Molecular Sciences
https://www.readbyqxmd.com/read/29383115/circulating-cd9-gfap-survivin-exosomes-in-malignant-glioma-patients-following-survivin-vaccination
#12
Phillip M Galbo, Michael J Ciesielski, Sheila Figel, Orla Maguire, Jingxin Qiu, Laura Wiltsie, Hans Minderman, Robert A Fenstermaker
Glioma cells release exosomes in culture and into the extracellular matrix in vivo. These nanobodies transport an array of biomolecules and are capable of mediating cell-cell communication. Circulating exosomes in cancer patients may be indicative of disease status and response to therapy. The inhibitor of apoptosis protein (IAP) survivin (SVN) promotes cancer cell proliferation, local immune suppression and resistance to chemotherapy and it is a potential cancer biomarker. We used imaging flow cytometry to perform quantitative measurements of circulating SVN+ exosomes in the serum of malignant glioma patients undergoing investigational treatment with an anti-survivin vaccine (SurVaxM)...
December 29, 2017: Oncotarget
https://www.readbyqxmd.com/read/29382529/homology-modeling-and-docking-of-aahii-nanobody-complexes-reveal-the-epitope-binding-site-on-aahii-scorpion-toxin
#13
Ayoub Ksouri, Kais Guedira, Rahma Ben Abderrazek, B A Gowri Shankar, Alia Ben Kahla, Ozlem Tanstan Bishop, Balkiss Bouhaouala-Zahar
Scorpion envenoming and its treatment is a public health problem in many parts of the world due to highly toxic venom polypeptides diffusing rapidly within the body of severely envenomed victims. Recently, 38 AahII-specific Nanobody sequences (Nbs) were retrieved from which the performance of NbAahII10 nanobody candidate, to neutralize the most poisonous venom compound namely AahII acting on sodium channels, was established. Herein, structural computational approach is conducted to elucidate the Nb-AahII interactions that support the biological characteristics, using Nb multiple sequence alignment (MSA) followed by modeling and molecular docking investigations (RosettaAntibody, ZDOCK software tools)...
January 27, 2018: Biochemical and Biophysical Research Communications
https://www.readbyqxmd.com/read/29382428/-construction-and-identification-of-nanobody-phage-display-library-targeting-middle-east-respiratory-syndrome-coronavirus
#14
Lei He, Jiangfan Li, Shuo Ren, Shihui Sun, Yan Guo, Hongjie Qiu, Yuanxiang Liao, Kaiyuan Ji, Ruiwen Fan, Guangyu Zhao, Yusen Zhou
Objective To construct a phage display library of specific nano-antibodies against the Middle East respiratory syndrome coronavirus (MERS-CoV) and apply it to the screening of neutralizing nano-antibodies. Methods MERS-CoV receptor-binding domain (RBD) recombinant protein was used to immunize alpaca. After the last immunization, peripheral blood mononuclear cells (PBMCs) were isolated from the whole blood and total RNA was extracted. The VHH gene was amplified by PCR and used to construct recombinant phages...
December 2017: Xi Bao Yu Fen Zi Mian Yi Xue za Zhi, Chinese Journal of Cellular and Molecular Immunology
https://www.readbyqxmd.com/read/29374062/protein-binders-and-their-applications-in-developmental-biology
#15
REVIEW
Stefan Harmansa, Markus Affolter
Developmental biology research would benefit greatly from tools that enable protein function to be regulated, both systematically and in a precise spatial and temporal manner, in vivo In recent years, functionalized protein binders have emerged as versatile tools that can be used to target and manipulate proteins. Such protein binders can be based on various scaffolds, such as nanobodies, designed ankyrin repeat proteins (DARPins) and monobodies, and can be used to block or perturb protein function in living cells...
January 26, 2018: Development
https://www.readbyqxmd.com/read/29353171/microcystin-lr-nanobody-screening-from-an-alpaca-phage-display-nanobody-library-and-its-expression-and-application
#16
Chongxin Xu, Ying Yang, Liwen Liu, Jianhong Li, Xiaoqin Liu, Xiao Zhang, Yuan Liu, Cunzheng Zhang, Xianjin Liu
Microcystin-LR (MC-LR) is a type of biotoxin that pollutes the ecological environment and food. The study aimed to obtain new nanobodies from phage nanobody library for determination of MC-LR. The toxin was conjugated to keyhole limpet haemocyanin (KLH) and bovine serum albumin (BSA), respectively, then the conjugates were used as coated antigens for enrichment (coated MC-LR-KLH) and screening (coated MC-LR-BSA) of MC-LR phage nanobodies from an alpaca phage display nanobody library. The antigen-specific phage particles were enriched effectively with four rounds of biopanning...
January 17, 2018: Ecotoxicology and Environmental Safety
https://www.readbyqxmd.com/read/29334397/recombinant-phosphatidylserine-binding-nanobodies-for-targeting-of-extracellular-vesicles-to-tumor-cells-a-plug-and-play-approach
#17
Sander A A Kooijmans, Jerney J J M Gitz-Francois, Raymond M Schiffelers, Pieter Vader
Extracellular vesicles (EVs) are increasingly being recognized as candidate drug delivery systems due to their ability to functionally transfer biological cargo between cells. However, manipulation of targeting properties of EVs through engineering of the producer cells can be challenging and time-consuming. As a novel approach to confer tumor targeting properties to isolated EVs, we generated recombinant fusion proteins of nanobodies against the epidermal growth factor receptor (EGFR) fused to phosphatidylserine (PS)-binding domains of lactadherin (C1C2)...
January 15, 2018: Nanoscale
https://www.readbyqxmd.com/read/29331148/isolation-of-anti-extra-cellular-vesicle-single-domain-antibodies-by-direct-panning-on-vesicle-enriched-fractions
#18
Milica Popovic, Elisa Mazzega, Barbara Toffoletto, Ario de Marco
BACKGROUND: The thorough understanding of the physiological and pathological processes mediated by extracellular vesicles (EVs) is challenged by purification methods which are cumbersome, not reproducible, or insufficient to yield homogeneous material. Chromatography based on both ion-exchange and immune-capture can represent an effective method to improve EV purification and successive analysis. METHODS: Cell culture supernatant was used as a model sample for assessing the capacity of anion-exchange chromatography to separate distinct EV fractions and to isolate nanobodies by direct panning on whole EVs to recover binders specific for the native conformation of EV-surface epitopes and suitable to develop EV immune-capture reagents...
January 13, 2018: Microbial Cell Factories
https://www.readbyqxmd.com/read/29328338/binding-affinity-prediction-of-nanobody-protein-complexes-by-scoring-of-molecular-dynamics-trajectories
#19
Miguel A Soler, Sara Fortuna, Ario de Marco, Alessandro Laio
Nanobodies offer a viable alternative to antibodies for engineering high affinity binders. Their small size has an additional advantage: it allows exploiting computational protocols for optimizing their biophysical features, such as the binding affinity. The efficient prediction of this quantity is still considered a daunting task especially for modelled complexes. We show how molecular dynamics can successfully assist in the binding affinity prediction of modelled nanobody-protein complexes. The approximate initial configurations obtained by in silico design must undergo large rearrangements before achieving a stable conformation, in which the binding affinity can be meaningfully estimated...
January 12, 2018: Physical Chemistry Chemical Physics: PCCP
https://www.readbyqxmd.com/read/29327239/a-small-protein-probe-for-correlated-microscopy-of-endogenous-proteins
#20
Marit A de Beer, Jeroen Kuipers, Paul M P van Bergen En Henegouwen, Ben N G Giepmans
Probes are essential to visualize proteins in their cellular environment, both using light microscopy as well as electron microscopy (EM). Correlated light microscopy and electron microscopy (CLEM) requires probes that can be imaged simultaneously by both optical and electron-dense signals. Existing combinatorial probes often have impaired efficiency, need ectopic expression as a fusion protein, or do not target endogenous proteins. Here, we present FLIPPER-bodies to label endogenous proteins for CLEM. Fluorescent Indicator and Peroxidase for Precipitation with EM Resolution (FLIPPER), the combination of a fluorescent protein and a peroxidase, is fused to a nanobody against a target of interest...
January 11, 2018: Histochemistry and Cell Biology
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