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antibody library

Angela Chiew Wen Ch'ng, Azimah Ahmad, Zoltán Konthur, Theam Soon Lim
Panning is a common process used for antibody selection from phage antibody libraries. There are several methods developed for a similar purpose, namely streptavidin mass spectrometry immunoassay (MSIA™) Disposable Automation Research Tips, magnetic beads, polystyrene immunotubes, and microtiter plate. The advantage of using a magnetic particle processor system is the ability to carry out phage display panning against multiple target antigens simultaneously in parallel. The system carries out the panning procedure using magnetic nanoparticles in microtiter plates...
2019: Methods in Molecular Biology
Viola Fühner, Philip Alexander Heine, Kilian Johannes Carl Zilkens, Doris Meier, Kristian Daniel Ralph Roth, Gustavo Marçal Schmidt Garcia Moreira, Michael Hust, Giulio Russo
Antibodies are widely used in a large variety of research applications, for diagnostics and therapy of numerous diseases, primarily cancer and autoimmune diseases. Antibodies are binding specifically to target structures (antigens). The antigen-binding properties are not only dependent on the antibody sequence, but also on the discrete antigen region recognized by the antibody (epitope). Knowing the epitope is valuable information for the improvement of diagnostic assays or therapeutic antibodies, as well as to understand the immune response of a vaccine...
2019: Methods in Molecular Biology
Fortunato Ferrara, Maria Felicia Soluri, Daniele Sblattero
In vitro display technologies have put together the generation of large antibody libraries with selection and screening procedures to identify lead candidates. Phage display antibody libraries allow selecting and identifying binders for a variety of antigens. Nonetheless, the procedure is limited by the possibility to quantitatively follow the enrichment during selection cycles and tune up the clones for specific binding proprieties (i.e., affinity). Yeast display allows the expression of thousands of copies of the antibody on each cell, simultaneously carrying the plasmid encoding that antibody, moreover the selection parameters can be accurately controlled by flow cytometry-based analysis and sorting...
2019: Methods in Molecular Biology
Marco Dal Ferro, Serena Rizzo, Emanuela Rizzo, Francesca Marano, Immacolata Luisi, Olga Tarasiuk, Daniele Sblattero
During the last 20 years in vitro technologies opened powerful routes to combine the generation of large libraries together with fast selection and screening procedures to identify lead candidates. One of the most successful methods is based on the use of filamentous phages. Functional Antibodies (Abs) fragments can be displayed on the surface of phages by fusing the coding sequence of the antibody variable (V) regions to the phage minor coat protein pIII. By creating large libraries, antibodies with affinities comparable to those obtained using traditional hybridoma technology can be isolated by a series of cycles of selection on the antigen of interest...
2019: Methods in Molecular Biology
Koji Hashimoto, Kohei Kurosawa, Hidetaka Seo, Kunihiro Ohta
We previously developed the in vitro method to generate monoclonal antibodies (mAbs) from libraries constructed with chicken B-cell line DT40 (referred to as the "ADLib system"). As the wild-type DT40 cells express immunoglobulin M (IgM), the original ADLib system provides monoclonal antibodies in chicken IgM format. For the therapeutic, diagnostic, and research purposes, the Fc regions of IgMs should be exchanged to other classes and species, for example human or murine IgG. However, the Fc engineering by conventional bioengineering process is laborious and takes plenty of time...
2019: Methods in Molecular Biology
Matthias Rüdt, Sebastian Andris, Robin Schiemer, Jürgen Hubbuch
Current biopharmaceutical production heavily relies on chromatography for protein purification. Recently, research has intensified towards finding suitable solutions to monitoring the chromatographic steps by multivariate spectroscopic sensors. Here, hard-constraint multivariate curve resolution (MCR) was investigated as a calibration-free method for factorizing bilinear preparative protein chromatograms into concentrations and spectra. Protein elutions were assumed to follow exponentially modified Gaussian (EMG) curves...
November 26, 2018: Journal of Chromatography. A
Qi Tang, Siping Xiong, Xudong Liang, Xingwang Kuai, Yiwen Wang, Changjun Wang, Zhenqing Feng, Jin Zhu
BACKGROUND: Disease caused by Bacillus anthracis is often accompanied by high mortality primarily due to toxin-mediated injury. In the early disease course, anthrax toxins are secreted; thus, antibiotic use is limited to the early stage. In this regard, antibodies against the toxin component, protective antigen (PA), play an important role in protecting against anthrax. Therefore, we developed PA21, a fully human anti-PA immunoglobulin G monoclonal antibody. METHODS: Combining human Fab was screened from a phage library with human heavy constant regions...
December 10, 2018: BMC Infectious Diseases
Jianglong Peng, Hao Yin, Ying Zhou, Haoyuan Jia, Yubao Cui
BACKGROUND: The dust mite Dermatophagoides farinae is a common worldwide cause of indoor allergies induced by mite proteins, including the mid-tier allergen Der f 7. Objective To identify conformational epitopes in Der f 7 using mimotope mapping and computational modelling. METHODS: Here, we used standard hybridoma technology to generate 3 new monoclonal antibodies against Der f 7 and performed mimotope mapping by probing a random peptide phage display library. Computational tools, including Minox and the DiscoTope-2...
December 8, 2018: Protein and Peptide Letters
Jhih-Wei Jian, Hong-Sen Chen, Yi-Kai Chiu, Hung-Pin Peng, Chao-Ping Tung, Ing-Chien Chen, Chung-Ming Yu, Yueh-Liang Tsou, Wei-Ying Kuo, Hung-Ju Hsu, An-Suei Yang
Antibodies provide immune protection by recognizing antigens of diverse chemical properties, but elucidating the amino acid sequence-function relationships underlying the specificity and affinity of antibody-antigen interactions remains challenging. We designed and constructed phage-displayed synthetic antibody libraries with enriched protein antigen-recognition propensities calculated with machine learning predictors, which indicated that the designed single-chain variable fragment variants were encoded with enhanced distributions of complementarity-determining region (CDR) hot spot residues with high protein antigen recognition propensities in comparison with those in the human antibody germline sequences...
December 7, 2018: MAbs
Dina Schneider, Ying Xiong, Peirong Hu, Darong Wu, Weizao Chen, Tianlei Ying, Zhongyu Zhu, Dimiter S Dimitrov, Boro Dropulic, Rimas J Orentas
Acute myeloid leukemia (AML) remains a challenging pediatric and adult disease. Given the elevated expression of the CD33 antigen on leukemic blasts, therapeutic approaches to AML now feature the approved antibody drug conjugate (Mylotarg, GO) and investigational CART cell approaches incorporating CD33-binding domains derived from humanized scFvs. We designed a functional chimeric antigen receptor utilizing a human targeting sequence, derived from a heavy chain variable domain, termed CAR33VH. Lentiviral-based expression vectors which encoded CAR constructs incorporating the novel binding domain (CAR33VH), or the My96 scFv control binder (My96CAR) in frame with a CD8 hinge and transmembrane domain, a 4-1BB costimulatory domain and a CD3 zeta activation domain, were transduced into primary human CD4+ and CD8+ T cells, and CAR expression was confirmed by flow cytometry...
2018: Frontiers in Oncology
Adriana-Michelle Wolf Pérez, Pietro Sormanni, Jonathan Sonne Andersen, Laila Ismail Sakhnini, Ileana Rodriguez-Leon, Jais Rose Bjelke, Annette Juhl Gajhede, Leonardo De Maria, Daniel E Otzen, Michele Vendruscolo, Nikolai Lorenzen
Despite major advances in antibody discovery technologies, the successful development of monoclonal antibodies (mAbs) into effective therapeutic and diagnostic agents can often be impeded by developability liabilities, such as poor expression, low solubility, high viscosity and aggregation. Therefore, strategies to predict at the early phases of antibody development the risk of late-stage failure of antibody candidates are highly valuable. In this work, we employ the in silico solubility predictor CamSol to design a library of 17 variants of a humanized mAb predicted to span a broad range of solubility values, and we examine their developability potential with a battery of commonly used in vitro and in silico assays...
December 7, 2018: MAbs
Jacob P Turowec, Esther W T Lau, Xiaowei Wang, Kevin R Brown, Frederic A Fellouse, Kamaldeep K Jawanda, James Pan, Jason Moffat, Sachdev Sidhu
Dysregulation of the ErbB family of receptor tyrosine kinases is involved in the progression of many cancers. Antibodies targeting the dimerization domains of family members EGFR and HER2 are approve cancer therapeutics, but efficacy is restricted to a subset of tumors and resistance often develops in response to treatment. A third family member, HER3, heterodimerizes with both EGFR and HER2 and has also been implicated in cancer. Consequently, there is strong interest in developing antibodies that target HER3, but to date, no therapeutics have been approved...
December 6, 2018: Journal of Biological Chemistry
Stefan Harth, Andre Ten Haaf, Christian Loew, Christian Frisch, Achim Knappik
Anti-idiotypic antibodies play an important role in pre-clinical and clinical development of therapeutic antibodies, where they are used for pharmacokinetic studies and for the development of immunogenicity assays. By using an antibody phage display library in combination with guided in vitro selection against various marketed drugs, we generated antibodies that recognize the drug only when bound to its target. We have named such specificities Type 3, to distinguish them from the anti-idiotypic antibodies that specifically detect free antibody drug or total drug...
December 5, 2018: MAbs
Peng-Cheng Yu, Xiao-Yan Tao, Li-Hua Wang, Qing Tang, Li-Yun Fan, Shu-Xia Zhang, Shu-Qing Liu, Xue-Xin Lu, Gui-Zhen Wu, Wu-Yang Zhu
BACKGROUND: The injection of rabies immune globulin (RIG) is of the utmost importance in the management of category III exposures to rabies-suspect animals. Because of the high cost and limited availability of existing RIG, one possible replacement for RIG is monoclonal antibodies (MAbs) against the rabies virus (RABV). Consequently, it is necessary to determine the neutralizing activity of the MAbs against rabies viruses, especially street rabies virus. However, the method to detect the neutralizing activity of MAbs against street rabies virus remains undefined...
December 5, 2018: Infectious Diseases of Poverty
Jirawat Khanongnoi, Siratcha Phanthong, Onrapak Reamtong, Anchalee Tungtronchitr, Wanpen Chaicumpa, Nitat Sookrung
Snake venom-metalloproteinases (SVMPs) are the primary factors that disturb hemostasis and cause hemorrhage in the venomous snake bitten subjects. Kaouthiagin is a unique SVMP that binds and cleaves von Willebrand factor (vWF) at a specific peptide bond leading to inhibition of platelet aggregation, which enhances the hemorrhage. Kaouthiagin is a low abundant venom component of Thai cobra ( Naja kaouthia ); thus, most horse-derived antivenins used for cobra bite treatment do not contain adequate anti-kaouthiagin...
December 3, 2018: Toxins
Christa Nederstigt, Bas Uitbeijerse, Laura Janssen, E P Corssmit, Eelco de Koning, Olaf Dekkers
INTRODUCTION: The association between type 1 diabetes (T1D) and other auto-immune diseases is well-known. However, a quantitative overview of all associated auto-immune diseases and their prevalence in T1D is lacking. METHODS: We searched Pubmed, Web of Science, EMBASE and Cochrane library in September 2018 to identify relevant articles about the prevalence of the following associated auto-immune diseases in T1D cohorts: auto-immune thyroid disease, celiac disease, gastric autoimmunity including pernicious anemia, vitiligo, and adrenal gland insufficiency...
December 1, 2018: European Journal of Endocrinology
Lutz Frönicke, Denise N Bronner, Mariana X Byndloss, Bridget McLaughlin, Andreas J Bäumler, Alexander J Westermann
Dual RNA-seq has emerged as a genome-wide expression profiling technique, simultaneously measuring RNA transcript levels in a given host and its pathogen during an infection. Recently, the method was transferred from cell culture to in vivo models of bacterial infections; however, specific host cell-type resolution has not yet been achieved. Here we present a detailed protocol that describes the application of Dual RNA-seq to murine colonocytes isolated from mice infected with the enteric pathogen Salmonella Typhimurium...
2018: Methods in Enzymology
Itoko Hayashi, Seiji Kanda, Pheophet Lamaningao, Nobuyuki Mishima, Toshimasa Nishiyama
This study focuses on the host-parasite relationship of human Ascaris lumbricoides, which is a parasite of the small intestine and is also one of the commonest parasites worldwide. As part of this investigation, we examined the host-parasite relationship assuming that there is a common antigenicity, shared protein between A. lumbricoides and human small intestinal mucosa, using molecular techniques. We obtained three DNA clones from human colon cDNA library by screening for anti-A. lumbricoides polyclonal antibodies...
November 24, 2018: Molecular and Biochemical Parasitology
Michael J Lowden, Kevin A Henry
Antibody (Ab) repertoire sequencing using high-throughput massively parallel technologies has contributed substantially to the understanding of Ab responses following infection, vaccination and autoimmunity. Because individual B-cell receptors are recombined and diversified somatically, genomic comparisons are limited, and distinguishing rare variants from sequencing errors is a major challenge. Oxford Nanopore Technologies' MinION is a highly portable and cost-effective third-generation sequencing instrument, but has not been used for Ab repertoire sequencing due to its high error rate (approximately 1/10 bases)...
December 2018: BioTechniques
Angela Bekesi, Sara Abdellaoui, Natalie Holroyd, Wouter Van Delm, Els Pardon, Jarne Pauwels, Kris Gevaert, Jan Steyaert, Stefaan Derveaux, Antoni Borysik, Peter Tompa
The structural and functional characterization of large multidomain signaling proteins containing long disordered linker regions represents special methodological and conceptual challenges. These proteins show extreme structural heterogeneity and have complex posttranslational modification patterns, due to which traditional structural biology techniques provide results that are often difficult to interpret. As demonstrated through the example of two such multidomain proteins, CREB-binding protein (CBP) and its paralogue, p300, even the expression and purification of such proteins are compromised by their extreme proteolytic sensitivity and structural heterogeneity...
2018: Methods in Enzymology
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