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https://www.readbyqxmd.com/read/28336958/from-rabbit-antibody-repertoires-to-rabbit-monoclonal-antibodies
#1
REVIEW
Justus Weber, Haiyong Peng, Christoph Rader
In this review, we explain why and how rabbit monoclonal antibodies have become outstanding reagents for laboratory research and increasingly for diagnostic and therapeutic applications. Starting with the unique ontogeny of rabbit B cells that affords highly distinctive antibody repertoires rich in in vivo pruned binders of high diversity, affinity and specificity, we describe the generation of rabbit monoclonal antibodies by hybridoma technology, phage display and alternative methods, along with an account of successful humanization strategies...
March 24, 2017: Experimental & Molecular Medicine
https://www.readbyqxmd.com/read/28304351/the-preparation-and-identification-of-a-monoclonal-antibody-against-citrinin-and-the-development-of-detection-via-indirect-competitive-elisa
#2
Shimuye Kalayu Yirga, Sumei Ling, Yanling Yang, Jun Yuan, Shihua Wang
Citrinin (CTN) is a hepato-nephrotoxic mycotoxin produced by fungi genera of Aspergillus, Monauscus, and Penicillium. CTN contaminates grains, fruits, juices and vegetables, and causes various toxic effects to humans and animals. It has small molecular weight, which is non-immunogenic to animals. Thus, CTN was conjugated to bovine serum albumin (BSA) and ovalbumin (OVA), respectively, by amide bonds using 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC) and N-hydroxysuccinimide (NHS). Mice were immunized with CTN-BSA conjugates, and spleen cells of the immunized mice were fused with Sp2/0 myeloma cells to obtain 21H27 hybriodoma cell...
March 17, 2017: Toxins
https://www.readbyqxmd.com/read/28296410/a-light-responsive-nanoparticle-based-delivery-system-using-pheophorbide-a-graft-polyethyleneimine-for-dendritic-cell-based-cancer-immunotherapy
#3
Chuangnian Zhang, Ju Zhang, Gaona Shi, Huijuan Song, Shengbin Shi, Xiuyuan Zhang, Pingsheng Huang, Zhihong Wang, Weiwei Wang, Chun Wang, Deling Kong, Chen Li
In this study, the photochemical internalization (PCI) technique was adopted in a nanoparticle-based antigen delivery system to enhance antigen-specific CD8(+) T cell immune response for cancer immunotherapy. Pheophorbide-A, a hydrophobic photosensitizer, grafted with polyethyleneimine (PheoA-PEI) with endosome escape activity and near infrared imaging capability was prepared. A model antigen ovalbumin (OVA) was then complexed with PheoA-PEI to form PheoA-PEI/OVA nanoparticles (PheoA-PEI/OVA NPs) that are responsive to light...
March 15, 2017: Molecular Pharmaceutics
https://www.readbyqxmd.com/read/28293253/can-anti-bothropstoxin-i-antibodies-discriminate-between-bothrops-jararaca-and-bothrops-jararacussu-venoms
#4
Ricardo Teixeira Araujo, Carlos Corrêa-Netto, Leonora Brazil-Más, Caio Raony Farina Silveira, Irene Fernandes, Russolina Benedeta Zingali
BACKGROUND: Snakes of the genus Bothrops, popularly known as pit vipers, are responsible for most cases of snakebite in Brazil. Within this genus, Bothrops jararacussu and B. jararaca deserve special attention due to the severity of their bites and for inhabiting densely populated areas. Regarding the treatment of snakebites by Bothrops jararacussu, questions have been raised about the effectiveness of the specific bothropic antivenom in neutralizing myotoxic effects; however, there are no accurate data for humans...
2017: Journal of Venomous Animals and Toxins Including Tropical Diseases
https://www.readbyqxmd.com/read/28286781/generation-of-monoclonal-antibodies-against-immunoglobulin-proteins-of-the-domestic-ferret-mustela-putorius-furo
#5
Greg A Kirchenbaum, Ted M Ross
The domestic ferret (Mustela putorius furo) serves as an animal model for the study of several viruses that cause human disease, most notably influenza. Despite the importance of this animal model, characterization of the immune response by flow cytometry (FCM) is severely hampered due to the limited number of commercially available reagents. To begin to address this unmet need and to facilitate more in-depth study of ferret B cells including the identification of antibody-secreting cells, eight unique murine monoclonal antibodies (mAb) with specificity for ferret immunoglobulin (Ig) were generated using conventional B cell hybridoma technology...
2017: Journal of Immunology Research
https://www.readbyqxmd.com/read/28282494/hapten-specific-single-cell-selection-of-hybridoma-clones-by-facs-for-the-generation-of-monoclonal-antibodies
#6
Martin Dippong, Peter Carl, Christine Lenz, Jörg Schenk, Katrin Hoffmann, Timm Schwaar, Rudolf J Schneider, Maren Kuhne
The conventional hybridoma screening and subcloning process is generally considered to be one of the most critical steps in hapten-specific antibody production. It is time-consuming, monoclonality is not guaranteed and the number of clones that can be screened is limited. Our approach employs a novel hapten-specific labeling technique of hybridoma cells. This allows for fluorescence-activated cell sorting (FACS) and single-cell deposition and thereby eliminating the abovementioned problems. A two-step staining approach is used to detect antigen-specificity and antibody expression: in order to detect antigen-specificity hybridoma cells are incubated with a hapten-peroxidase conjugate (hapten-HRP), which is subsequently incubated with a fluorophore-labeled polyclonal anti-peroxidase antibody (anti-HRP-Alexa Fluor® 488)...
March 10, 2017: Analytical Chemistry
https://www.readbyqxmd.com/read/28274321/-preparation-of-monoclonal-antibody-against-4-amylphenol-and-homology-modeling-of-its-fv-fragment
#7
Lei Cheng, Haizhen Wu, Jing Fei, Lujia Zhang, Jiang Ye, Huizhan Zhang
Objective To prepare and characterize a monoclonal antibody (mAb) against 4-amylphenol (4-AP), clone its cDNA sequence and make homology modeling for its Fv fragment. Methods A high-affinity anti-4-AP mAb was generated from a hybridoma cell line F10 using electrofusion between splenocytes from APA-BSA-immunized mouse and Sp2/0 myeloma cells. Then we extracted the mRNA of F10 cells and cloned the cDNA of mAb. The homology modeling and molecular docking of its Fv fragment was conducted with biological software...
March 2017: Xi Bao Yu Fen Zi Mian Yi Xue za Zhi, Chinese Journal of Cellular and Molecular Immunology
https://www.readbyqxmd.com/read/28269763/development-of-high-affinity-monoclonal-antibody-using-cd44-overexpressed-cells-as-a-candidate-for-targeted-immunotherapy-and-diagnosis-of-acute-myeloid-leukemia
#8
Amir Amanzadeh, Fatemeh Heidarnejad, Meghdad Abdollahpour-Alitappeh, Vahid Molla Kazemiha, Shamsi Yari, Ali Reza Hadizadeh Tasbiti, Mahdi Habibi Anbouhi, Mohsen Abolhassani, Mohammad Ali Shokrgozar
AIM: CD44s antigens have been suggested as an efficient biomarker for cancer stem cells. Current study aimed to develop a hybridoma that producing a high affinity murine anti-human CD44 monoclonal antibody for early diagnostic laboratory tests of some cancer. MATERIALS AND METHODS: To make hybridoma against CD44, mice were immunized with MDA-MB-468 cells. Resulted hybridomas using three culture media were screened by indirect ELISA, then cloned by limiting dilution, and isotype was determined after obtaining ascitic fluid and antibody purification...
February 25, 2017: Human Antibodies
https://www.readbyqxmd.com/read/28269762/cloning-and-molecular-characterization-of-the-cdnas-encoding-the-variable-regions-of-an-anti-cd20-monoclonal-antibody
#9
Dariush Shanehbandi, Jafar Majidi, Tohid Kazemi, Behzad Baradaran, Leili Aghebati-Maleki
BACKGROUND: CD20-based targeting of B-cells in hematologic malignancies and autoimmune disorders is associated with outstanding clinical outcomes. Isolation and characterization of VH and VL cDNAs encoding the variable regions of the heavy and light chains of monoclonal antibodies (MAb) is necessary to produce next generation MAbs and their derivatives such as and bispecific antibodies (bsAb) and single-chain variable fragments (scFv). OBJECTIVE: This study was aimed at cloning and characterization of the VH and VL cDNAs from a hybridoma cell against the CD20 antigen...
February 24, 2017: Human Antibodies
https://www.readbyqxmd.com/read/28239619/evaluation-of-cross-presentation-in-bone-marrow-derived-dendritic-cells-in-vitro-and-splenic-dendritic-cells-ex-vivo-using-antigen-coated-beads
#10
Andrés Alloatti, Fiorella Kotsias, Eik Hoffmann, Sebastian Amigorena
Antigen presentation by MHC class I molecules, also referred to as cross-presentation, elicits cytotoxic immune responses. In particular, dendritic cells (DC) are the most proficient cross-presenting cells, since they have developed unique means to control phagocytic and degradative pathways. This protocol allows the evaluation of antigen cross-presentation both in vitro (by using bone marrow-derived DC) and ex vivo (by purifying CD8(+) DC from spleen after incorporation of particulate antigen) using ovalbumin (OVA)-coupled particles...
November 20, 2016: Bio-protocol
https://www.readbyqxmd.com/read/28219820/versatile-carboxyl-terminus-of-capsid-protein-of-porcine-circovirus-type-2-were-recognized-by-monoclonal-antibodies-with-pluripotency-of-binding
#11
Ling-Chu Hung, Ivan-Chen Cheng
We designed the peptide (C3) mimetic carboxyl-terminus (Cterminus) of capsid protein of porcine circovirus type 2b (PCV2b-1A/1B) inducing humoral immunity and generating hybridomas. The positive reactivity of the mAbs to PCV2 capsid protein was demonstrated by Western blot assay. Those mAbs also showed positive signals on PCV2b infected swine lymphocytes by indirect immunofluorescence staining. The mAb 1H3 bound to three minimal linear epitopes (P62, DPPLNP; P67, DPPLNPK; P73, LKDPPLKP), which was located at Cterminus of the capsid protein of PCV2b-1A/1B, PCV2b-1C, and PCV2a-2A respectively...
February 17, 2017: Molecular Immunology
https://www.readbyqxmd.com/read/28216237/development-of-an-anti-ferret-cd4-monoclonal-antibody-for-the-characterisation-of-ferret-t-lymphocytes
#12
Daniel S Layton, Xiaowen Xiao, John D Bentley, Louis Lu, Cameron R Stewart, Andrew G D Bean, Timothy E Adams
The ferret is an established animal model for a number of human respiratory viral infections, such as influenza virus and more recently, Ebola virus. However, a paucity of immunological reagents has hampered the study of cellular immune responses. Here we describe the development and characterisation of a novel monoclonal antibody (mAb) against the ferret CD4 antigen and the characterisation of ferret CD4 T lymphocytes. Recombinant production and purification of the ferret CD4 ectodomain soluble protein allowed hybridoma generation and the generation of a mAb (FeCD4) showing strong binding to ferret CD4 protein and lymphoid cells by flow cytometry...
February 16, 2017: Journal of Immunological Methods
https://www.readbyqxmd.com/read/28186433/production-of-monoclonal-antibody-against-excretory-secretory-antigen-of-fasciola-hepatica-and-evaluation-of-its-efficacy-in-the-diagnosis-of-fascioliasis
#13
Samaneh Abdolahi Khabisi, Bahador Sarkari, Abdolali Moshfe, Sedigheh Jalali
Parasitological methods are not helpful for the diagnosis of fascioliasis in acute and invasive periods of the disease. Detection of coproantigens seems to be a suitable alternative approach in the diagnosis of fascioliasis. The present study aimed to develop a reliable antigen detection system, using monoclonal antibodies raised against excretory-secretory (ES) antigen of Fasciola hepatica, for the diagnosis of fascioliasis. Fasciola adult worms were collected from the bile ducts of infected animals. Species of the fluke was determined by polymerase chain reaction-restriction fragment length polymorphism (RFLP-PCR)...
February 2017: Monoclonal Antibodies in Immunodiagnosis and Immunotherapy
https://www.readbyqxmd.com/read/28154257/antibody-fragments-for-on-site-testing-of-cannabinoids-generated-via-in-vitro-affinity-maturation
#14
Izumi Morita, Hiroyuki Oyama, Mayumi Yasuo, Kazuhisa Matsuda, Kengo Katagi, Aya Ito, Hiroka Tatsuda, Hiroyuki Tanaka, Satoshi Morimoto, Norihiro Kobayashi
Law enforcement against illicit use of cannabis and related substances requires rapid, feasible, and reliable tools for on-site testing of cannabinoids. Notably, methods based on cannabinoid-specific antibodies enable efficient screening of multiple specimens. Antibody engineering may accelerate development of modern and robust testing systems. Here, we used in vitro affinity maturation to generate a single-chain Fv fragment (scFv) that recognizes with high affinity the psychoactive cannabinoid, Δ(9)-tetrahydrocannabinol (THC)...
2017: Biological & Pharmaceutical Bulletin
https://www.readbyqxmd.com/read/28144646/an-open-sandwich-immunoassay-for-detection-of-13-r-s-hydroxy-9-e-11-e-octadecadienoic-acid
#15
Jinhua Dong, Mototada Shichiri, Chan-I Chung, Takahiro Shibata, Koji Uchida, Yoshihisa Hagihara, Yasukazu Yoshida, Hiroshi Ueda
Lipid peroxidation is involved in many disorders and diseases such as cardiovascular disease, cancers, neurodegenerative diseases, and even aging. Lipid peroxidation products existing in blood or bodily fluids are very important biomarkers for the diagnosis of such diseases. In particular, 13(R,S)-hydroxy-9(E),11(E)-octadecadienoic acid (13-(E,E)-HODE) is an oxidiation product of linoleic acid, which is an important biomarker for many diseases such as diabetes and Alzheimer's disease. In this study, we successfully displayed the antigen-binding fragment of an antibody produced by hybridoma 1213-1 on the M13 phage and performed analysis of the antibody variable region genes...
February 1, 2017: Analyst
https://www.readbyqxmd.com/read/28137617/bioprocess-optimization-under-uncertainty-using-ensemble-modeling
#16
Yang Liu, Rudiyanto Gunawan
The performance of model-based bioprocess optimizations depends on the accuracy of the mathematical model. However, models of bioprocesses often have large uncertainty due to the lack of model identifiability. In the presence of such uncertainty, process optimizations that rely on the predictions of a single "best fit" model, e.g. the model resulting from a maximum likelihood parameter estimation using the available process data, may perform poorly in real life. In this study, we employed ensemble modeling to account for model uncertainty in bioprocess optimization...
February 20, 2017: Journal of Biotechnology
https://www.readbyqxmd.com/read/28125314/bisfabs-tools-for-rapidly-screening-hybridoma-iggs-for-their-activities-as-bispecific-antibodies
#17
Sanket Patke, Ji Li, Peiyin Wang, Dion Slaga, Jennifer Johnston, Sunil Bhakta, Siler Panowski, Liping L Sun, Teemu Junttila, Justin M Scheer, Diego A Ellerman
Bispecific antibodies are a growing class of therapeutic molecules. Many of the current bispecific formats require DNA engineering to convert the parental monoclonal antibodies into the final bispecific molecules. We describe here a method to generate bispecific molecules from hybridoma IgGs in 3-4 days using chemical conjugation of antigen-binding fragments (Fabs) (bisFabs). Proteolytic digestion conditions for each IgG isotype were analyzed to optimize the yield and quality of the final conjugates. The resulting bisFabs showed no significant amounts of homodimers or aggregates...
January 26, 2017: MAbs
https://www.readbyqxmd.com/read/28122974/monoclonal-antibodies-to-v2-v3-the-cd4-binding-site-and-gp41-hiv-1-mediate-phagocytosis-in-a-dose-dependent-manner
#18
Thomas Musich, Liuzhe Li, Lily Liu, Susan Zolla-Pazner, Marjorie Robert-Guroff, Miroslaw K Gorny
In light of weak or absent neutralizing activity mediated by anti-V2 monoclonal Abs (mAbs), we tested whether they can mediate Ab-dependent cellular phagocytosis (ADCP) which is an important element of anti-HIV-1 immunity. We tested six anti-V2 mAbs and compared them with 21 mAbs specific for V3, the CD4-binding site (CD4bs), and gp41 derived from HIV-1 chronically infected individuals and produced by hybridoma cells. ADCP activity was measured by flow cytometry using uptake by THP-1 monocytic cells of fluorescent beads coated with gp120, gp41, BG505 SOSIP...
January 25, 2017: Journal of Virology
https://www.readbyqxmd.com/read/28119207/epitope-dependent-mechanisms-of-cd27-neutralization-revealed-by-x-ray-crystallography
#19
Galina Obmolova, Alexey Teplyakov, Thomas J Malia, Nicole Wunderler, Deborah Kwok, Linda Barone, Raymond Sweet, Tatiana Ort, Michael Scully, Gary L Gilliland
CD27 is a T and B cell co-stimulatory protein of the TNF receptor superfamily dependent on the availability of the TNF-like ligand CD70. Two anti-CD27 neutralizing monoclonal antibodies were obtained from mouse hybridoma and subsequently humanized and optimized for binding the target. The two antibodies are similar in terms of their CD27-binding affinity and ability to block NF-κB signaling, however their clearance rates in monkeys are very different. The pharmacokinetics profiles could be epitope dependent...
March 2017: Molecular Immunology
https://www.readbyqxmd.com/read/28117810/generation-of-murine-monoclonal-antibodies-by-hybridoma-technology
#20
Pamela Holzlöhner, Katja Hanack
Monoclonal antibodies are universal binding molecules and are widely used in biomedicine and research. Nevertheless, the generation of these binding molecules is time-consuming and laborious due to the complicated handling and lack of alternatives. The aim of this protocol is to provide one standard method for the generation of monoclonal antibodies using hybridoma technology. This technology combines two steps. Step 1 is an appropriate immunization of the animal and step 2 is the fusion of B lymphocytes with immortal myeloma cells in order to generate hybrids possessing both parental functions, such as the production of antibody molecules and immortality...
January 2, 2017: Journal of Visualized Experiments: JoVE
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