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https://www.readbyqxmd.com/read/28444967/co-delivery-of-microrna-21-antisense-oligonucleotides-and-gemcitabine-using-nanomedicine-for-pancreatic-cancer-therapy
#1
Yaqing Li, Yinting Chen, Jiajia Li, Zuoquan Zhang, Chumei Huang, Guoda Lian, Kege Yang, Shaojie Chen, Ying Lin, Lingyun Wang, Kaihong Huang, Linjuan Zeng
Tumor metastasis occurs naturally in pancreatic cancer, and the efficacy of chemotherapy is usually poor. Precision medicine, combining down-regulation of target genes with chemotherapy drugs, is expected to improve the therapeutic effects. Therefore, we developed a combined therapy of microRNA-21 antisense oligonucleotides (ASO-miR-21) and Gemcitabine (Gem) using a targeted co-delivery nanoparticle (NP) carrier and investigated the synergistic inhibitory effects on pancreatic cancer cells metastasis and growth...
April 26, 2017: Cancer Science
https://www.readbyqxmd.com/read/28444391/dual-display-phage-selection-driven-by-co-engagement-of-two-targets-by-two-different-antibody-fragments
#2
Séverine Fagète, Ledicia Botas-Perez, Irène Rossito-Borlat, Kenneth Adea, Franck Gueneau, Ulla Ravn, François Rousseau, Marie Kosco-Vilbois, Nicolas Fischer, Oliver Hartley
Antibody phage display technology has supported the emergence of numerous therapeutic antibodies. The development of bispecific antibodies, a promising new frontier in antibody therapy, could be facilitated by new phage display approaches that enable pairs of antibodies to be co-selected based on co-engagement of their respective targets. We describe such an approach, making use of two complementary leucine zipper domains that heterodimerize with high affinity. Phagemids encoding a first antibody fragment (scFv) fused to phage coat protein via the first leucine zipper are rescued in bacteria expressing a second scFv fused to the second leucine zipper as a soluble periplasmic protein, so that it is acquired by phage during assembly...
April 24, 2017: Protein Engineering, Design & Selection: PEDS
https://www.readbyqxmd.com/read/28425967/a-simple-and-specific-noncompetitive-elisa-method-for-ht-2-toxin-detection
#3
Henri O Arola, Antti Tullila, Alexis V Nathanail, Tarja K Nevanen
We developed an HT-2 toxin-specific simple ELISA format with a positive read-out. The assay is based on an anti-immune complex (IC) scFv antibody fragment, which is genetically fused with alkaline phosphatase (AP). The anti-IC antibody specifically recognizes the IC between a primary anti-HT-2 toxin Fab fragment and an HT-2 toxin molecule. In the IC ELISA format, the sample is added together with the scFv-AP antibody to the ELISA plate coated with the primary antibody. After 15 min of incubation and a washing step, the ELISA response is read...
April 20, 2017: Toxins
https://www.readbyqxmd.com/read/28423625/inhibition-activity-of-a-disulfide-stabilized-diabody-against-basic-fibroblast-growth-factor-in-lung-cancer
#4
Yaxiong Cai, Shuange Yao, Jiangchuan Zhong, Jinxia Zhang, Haowu Jiang, Yanrui Deng, Ning Deng
The over-expression of basic fibroblast growth factor (bFGF) plays a crucial role in the development, invasion and metastasis of lung cancer. Therefore, neutralizing antibodies against bFGF may inhibit the growth of lung cancer. In this study, a Disulfide-stabilized diabody (ds-Diabody) against bFGF was constructed by site-directed mutation and overlap extension PCR (SOE-PCR) at the position of VH44 and VL100 in the scFv. The ds-Diabody was constructed and expressed in Pichia pastoris. We found that the ds-Diabody against bFGF could efficiently suppress the proliferation, migration and invasion of human lung cancer A549 cells in vitro...
March 21, 2017: Oncotarget
https://www.readbyqxmd.com/read/28423057/insertion-of-a-ligand-to-her2-in-gb-retargets-hsv-tropism-and-obviates-the-need-for-activation-of-the-other-entry-glycoproteins
#5
Biljana Petrovic, Tatiana Gianni, Valentina Gatta, Gabriella Campadelli-Fiume
Herpes simplex virus (HSV) entry into the cells requires glycoproteins gD, gH/gL and gB, activated in a cascade fashion by conformational modifications induced by cognate receptors and intermolecular signaling. The receptors are nectin1 and HVEM (Herpes virus entry mediator) for gD, and αvβ6 or αvβ8 integrin for gH. In earlier work, insertion of a single chain antibody (scFv) to the cancer receptor HER2 (human epidermal growth factor receptor 2) in gD, or in gH, resulted in HSVs specifically retargeted to the HER2-positive cancer cells, hence in highly specific non-attenuated oncolytic agents...
April 19, 2017: PLoS Pathogens
https://www.readbyqxmd.com/read/28421663/cloning-and-plant-based-production-of-antibody-mc10e7-for-a-lateral-flow-immunoassay-to-detect-4-arginine-microcystin-in-fresh-water
#6
Stanislav Melnik, A-C Neumann, R Karongo, S Dirndorfer, Martin Stübler, Verena Ibl, R Niessner, Dietmar Knopp, Eva Stoger
Antibody MC10E7 is one of a small number of monoclonal antibodies that bind specifically to [Arg4]-microcystins and it can be used to survey natural water sources and food samples for algal toxin contamination. However, the development of sensitive immunoassays in different test formats, particularly user-friendly tests for on-site analysis, requires a sensitive but also cost-effective antibody. The original version of MC10E7 was derived from a murine hybridoma, but we determined the sequence of the variable regions using the peptide mass-assisted cloning strategy and expressed a scFv (single-chain variable fragment) format of this antibody in yeast and a chimeric full size version in leaves of Nicotiana tabacum and N...
April 19, 2017: Plant Biotechnology Journal
https://www.readbyqxmd.com/read/28419225/epitope-mapping-of-a-new-anti-tn-antibody-detecting-gastric-cancer-cells
#7
Nina Persson, Nicolai Stuhr-Hansen, Christian Risinger, Stefan Mereiter, António Polónia, Karol Polom, András Kovács, Franco Roviello, Celso Reis, Charlotte Welinder, Lena Danielsson, Bo Jansson, Ola Blixt
Here we introduce a novel scFv antibody, G2-D11, specific for two adjacent Tn-antigens (GalNAc -Ser/Thr) binding equally to three dimeric forms of the epitope, Ser-Thr, Thr-Thr and Thr-Ser. Compared to other anti-Tn reagents, the binding of G2-D11 is minimally influenced by the peptide structure, which indicates a high degree of carbohydrate epitope dominance and a low influence from the protein backbone. With a high affinity (KDapp = 1.3 × 10-8M) and no cross-reactivity to either sialyl-Tn epitope or blood group A antigens, scFv G2-D11 is an excellent candidate for a well-defined anti-Tn-antigen reagent...
April 13, 2017: Glycobiology
https://www.readbyqxmd.com/read/28398709/continuous-desalting-of-refolded-protein-solution-improves-capturing-in-ion-exchange-chromatography-a-seamless-process
#8
Nicole Walch, Alois Jungbauer
Truly continuous biomanufacturing processes enable an uninterrupted feed stream throughout the whole production without the need for holding tanks. We have utilized microporous anion and cation exchangers into which only salts, but not proteins, can penetrate into the pores for desalting of protein solutions, while diafiltration or dilution is usually employed for feed adjustments. Anion exchange and cation exchange chromatography columns were connected in series to remove both anions and cations. To increase operation performance, a continuous process was developed comprised of four columns...
April 11, 2017: Biotechnology Journal
https://www.readbyqxmd.com/read/28393196/preparation-of-human-single-chain-variable-fragment-antibodies-against-anaplastic-thyroid-carcinoma-and-single-photon-emission%C3%A2-computed-tomography-computed-tomography-imaging-in-tumor-bearing-nude-mice
#9
Qian Liu, Hua Pang, Qiong Liu, Jing Zhou, Ying Liu
Anaplastic thyroid carcinoma (ATC) is the most aggressive malignant thyroid tumor with the worst prognosis, and the response to treatment is poor. We investigated soluble human single-chain variable fragment (scFv) which provides unique information for diagnostics, and for monitoring and optimizing responses to therapy. Enzyme-linked immunosorbent assay (ELISA) was used to detect the expression of scFv. The expression and relative molecular mass of soluble scFv were assessed by sodium dodecyl sulfate‑polyacrylamide gel electrophoresis (SDS-PAGE) and western blotting, respectively...
April 5, 2017: Oncology Reports
https://www.readbyqxmd.com/read/28392906/selection-of-single-chain-variable-fragments-specific-for-mycobacterium-tuberculosis-esat-6-antigen-using-ribosome-display
#10
Shahrzad Ahangarzadeh, Mojgan Bandehpour, Bahram Kazemi
OBJECTIVES: Tuberculosis (TB) is still one of the problematic infectious diseases in developing countries, especially in Iran. In the present study, we applied ribosome display technique to select single chain variable fragments (scFvs) specific for the 6-kDa early secretory antigenic target (ESAT-6) antigen of Mycobacterium tuberculosis from a mouse scFv library. MATERIALS AND METHODS: The gene encoding ESAT-6 was cloned into pET22b(+) plasmid and expressed in Escherichia coli BL21 (DE3)...
March 2017: Iranian Journal of Basic Medical Sciences
https://www.readbyqxmd.com/read/28392422/the-effect-of-varying-the-peptide-linker-length-in-a-single-chain-variable-fragment-antibody-against-wogonin-glucuronide
#11
Madan Kumar Paudel, Seiichi Sakamoto, Le Van Huy, Hiroyuki Tanaka, Tomofumi Miyamoto, Satoshi Morimoto
Peptide linkers of three different lengths were constructed to join the variable regions of the heavy chain (VH) and the light chain (VL) in a single-chain variable fragment antibody (scFv) specific for wogonin glucuronide (Wgn) that has the structure VH-(GGGGS)n-VL (n=3, 5, or 7). The scFv antibodies, which were expressed in Escherichia coli, were derived from an anti-Wgn monoclonal antibody (315A). An indirect competitive enzyme-linked immunosorbent assay (icELISA) was used to evaluate their reactivity and sensitivity, which is also used for quantitative analysis of Wgn...
April 7, 2017: Journal of Biotechnology
https://www.readbyqxmd.com/read/28389318/engineering-of-a-novel-zipfv-using-leucine-zipper-motif-against-rabies-virus-glycoprotein-g-with-improved-protection-potency-in-vivo
#12
Hualong Xi, Kaixin Zhang, Yanchun Yin, Tiejun Gu, Qing Sun, Zhuang Li, Yue Cheng, Chunlai Jiang, Wei Kong, Yongge Wu
Rabies is an acute zoonotic infectious disease with a high fatality rate but is preventable with vaccination and rabies immunoglobulin (RIG). The single-chain Fv fragment (scFv), a small engineered antigen-binding protein derived from antibody variable heavy (VH) and light (VL) chains connected by a peptide linker, can potentially be used to replace RIG. Here, we produced two peptides VH-JUN-HIS and VL-FOS-HA separately in Escherichia coli and assembled them to form zipFv successfully in vitro. The new zipFv utilizes FOS and JUN leucine zippers to form an antibody structure similar to the IgG counterpart with two free N-terminal ends of VH and VL...
April 4, 2017: Immunology Letters
https://www.readbyqxmd.com/read/28388565/prokaryotic-expression-of-mlaa-34-and-generation-of-a-novel-human-scfv-against-mlaa-34-by-phage-display-technology
#13
Yang Zhang, Pengyu Zhang, Aili He, Yun Yang, Jianli Wang, Hui Zhang, Wanggang Zhang
MLAA-34 is a newly identified monocytic leukemia-associated antigen that is overexpressed in acute monocytic leukemia specifically, thus providing a novel target for the therapy of acute monocytic leukemia. In this study, we first expressed MLAA-34 protein in Escherichia coli (E.coli) BL21 (DE3) cells and purified it by nickel ion affinity chromatography with high purity (>90%). Then, MLAA-34 was used as antigen for biopanning anti-MLAA-34 single chain antibody fragment (ScFv) from a fully human ScFv library, and a high affinity ScFv named MA1 was selected by phage-ELISA...
March 28, 2017: Oncotarget
https://www.readbyqxmd.com/read/28363519/pretargeting-with-bispecific-fusion-proteins-facilitates-delivery-of-nanoparticles-to-tumor-cells-with-distinct-surface-antigens
#14
Qi Yang, Christina L Parker, Yukang Lin, Oliver W Press, Steven I Park, Samuel K Lai
Tumor heterogeneity, which describes the genetically and phenotypically distinct subpopulations of tumor cells present within the same tumor or patient, presents a major challenge to targeted delivery of diagnostic and/or therapeutic agents. An ideal targeting strategy should deliver a given nanocarrier to the full diversity of cancer cells, which is difficult to achieve with conventional ligand-conjugated nanoparticles. We evaluated pretargeting (i.e., multistep targeting) as a strategy to facilitate nanoparticle delivery to multiple target cells by measuring the uptake of biotinylated nanoparticles by lymphoma cells with distinct surface antigens pretreated with different bispecific streptavidin-scFv fusion proteins...
March 29, 2017: Journal of Controlled Release: Official Journal of the Controlled Release Society
https://www.readbyqxmd.com/read/28346811/improved-soluble-scfv-elisa-screening-approach-for-antibody-discovery-using-phage-display-technology
#15
Mohammad R Tohidkia, Maryam Sepehri, Shirin Khajeh, Jaleh Barar, Yadollah Omidi
Phage display technology (PDT) is a powerful tool for the isolation of recombinant antibody (Ab) fragments. Using PDT, target molecule-specific phage-Ab clones are enriched through the "biopanning" process. The individual specific binders are screened by the monoclonal scFv enzyme-linked immunosorbent assay (ELISA) that may associate with inevitable false-negative results. Thus, in this study, three strategies were investigated for optimization of the scFvs screening using Tomlinson I and J libraries, including (1) optimizing the expression of functional scFvs, (2) improving the sensitivity of ELISA, and (3) preparing different samples containing scFvs...
March 1, 2017: SLAS Discovery
https://www.readbyqxmd.com/read/28343398/an-anti-programmed-death-1-antibody-%C3%AE-pd-1-fusion-protein-that-self-assembles-into-a-multivalent-and-functional-%C3%AE-pd-1-nanoparticle
#16
Peng Zhao, Djordje Atanackovic, Shuyun Dong, Hideo Yagita, Xiao He, Mingnan Chen
Cancer immune checkpoint therapy has achieved remarkable clinical successes in various cancers. However, current immune checkpoint inhibitors block the checkpoint of not only the immune cells that are important to cancer therapy but also the immune cells that are irrelevant to the therapy. Such an indiscriminate blockade limits the efficacy and causes the autoimmune toxicity of the therapy. It might be beneficial to use a carrier to target immune checkpoint inhibitors to cancer-reactive immune cells. Here, we explore a method to load the inhibitors into carriers...
April 19, 2017: Molecular Pharmaceutics
https://www.readbyqxmd.com/read/28342851/site-specific-photocoupling-of-pbpa-mutated-scfv-antibodies-for-use-in-affinity-proteomics
#17
Mattias Brofelth, Lars Wagner Städe, Anna Isinger Ekstrand, Linn Petersson Edfeldt, Rebeka Kovačič, Thorbjørn Terndrup Nielsen, Kim Lambertsen Larsen, Laurent Duroux, Christer Wingren
Recombinant antibody libraries can provide a source of renewable and high-performing binders tailored for use in affinity proteomics. In this context, the process of generating site-specific 1:1 tagging/functionalization and/or orientated surface immobilization of antibodies has, however, proved to be challenging. Hence, novel ways of generating such engineered antibodies for use in affinity proteomics could have a major impact on array performance. In this study, we have further tailored the design of human recombinant scFv antibodies for site-specific photocoupling through the use of an unnatural amino acid (UAA) and the Dock'n'Flash technology...
March 22, 2017: Biochimica et Biophysica Acta
https://www.readbyqxmd.com/read/28340507/understanding-the-contribution-of-disulphide-bridges-to-the-folding-and-misfolding-of-an-anti-a%C3%AE-scfv
#18
Laia Montoliu-Gaya, Jose C Martínez, Sandra Villegas
ScFv-h3D6 is a single chain variable fragment that precludes Aβ peptide-induced cytotoxicity by withdrawing Aβ oligomers from the amyloid pathway to the worm-like pathway. Production of scFv molecules is not a straightforward procedure because of the occurrence of disulphide scrambled conformations generated in the refolding process. Here, we separately removed the disulphide bond of each domain and solved the scrambling problem; and then, we intended to compensate the loss of thermodynamic stability by adding three C-terminal elongation mutations previously described to stabilize the native fold of scFv-h3D6...
March 24, 2017: Protein Science: a Publication of the Protein Society
https://www.readbyqxmd.com/read/28336957/next-generation-sequencing-enables-the-discovery-of-more-diverse-positive-clones-from-a-phage-displayed-antibody-library
#19
Wonjun Yang, Aerin Yoon, Sanghoon Lee, Soohyun Kim, Jungwon Han, Junho Chung
Phage display technology provides a powerful tool to screen a library for a binding molecule via an enrichment process. It has been adopted as a critical technology in the development of therapeutic antibodies. However, a major drawback of phage display technology is that because the degree of the enrichment cannot be controlled during the bio-panning process, it frequently results in a limited number of clones. In this study, we applied next-generation sequencing (NGS) to screen clones from a library and determine whether a greater number of clones can be identified using NGS than using conventional methods...
March 24, 2017: Experimental & Molecular Medicine
https://www.readbyqxmd.com/read/28332627/cross-neutralizing-anti-hiv-1-human-single-chain-variable-fragments-scfvs-against-cd4-binding-site-and-n332-glycan-identified-from-a-recombinant-phage-library
#20
Lubina Khan, Rajesh Kumar, Ramachandran Thiruvengadam, Hilal Ahmad Parray, Muzamil Ashraf Makhdoomi, Sanjeev Kumar, Heena Aggarwal, Madhav Mohata, Abdul Wahid Hussain, Raksha Das, Raghavan Varadarajan, Jayanta Bhattacharya, Madhu Vajpayee, K G Murugavel, Suniti Solomon, Subrata Sinha, Kalpana Luthra
More than 50% of HIV-1 infection globally is caused by subtype_C viruses. Majority of the broadly neutralizing antibodies (bnAbs) targeting HIV-1 have been isolated from non-subtype_C infected donors. Mapping the epitope specificities of bnAbs provides useful information for vaccine design. Recombinant antibody technology enables generation of a large repertoire of monoclonals with diverse specificities. We constructed a phage recombinant single chain variable fragment (scFv) library with a diversity of 7.8 × 10(8) clones, using a novel strategy of pooling peripheral blood mononuclear cells (PBMCs) of six select HIV-1 chronically infected Indian donors whose plasma antibodies exhibited potent cross neutralization efficiency...
March 23, 2017: Scientific Reports
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