scholarly journals Engineered resistance to Nosema bombycis by in vitro expression of a single-chain antibody in Sf9-III cells

PLoS ONE ◽  
2018 ◽  
Vol 13 (2) ◽  
pp. e0193065 ◽  
Author(s):  
Yukang Huang ◽  
Jie Chen ◽  
Bin Sun ◽  
Rong Zheng ◽  
Boning Li ◽  
...  
Keyword(s):  
Single Chain ◽  
Single Chain Antibody ◽  
In Vitro Expression ◽  
Nosema Bombycis ◽  
Engineered Resistance

Single-chain Antibody Against Reg4 Suppresses Gastric Cancer Cell Growth and Enhances 5-FU-induced Cell Death in vitro

2019 ◽  
Vol 19 (5) ◽  
pp. 610-619 ◽  
Author(s):  
Xue-Qing Zhang ◽  
Lu-Ting Yu ◽  
Pei Du ◽  
Tian-Qi Yin ◽  
Zhi-Yuan Zhang ◽  
...  
Keyword(s):  
Gastric Cancer ◽  
Cell Proliferation ◽  
Cell Death ◽  
Cancer Cell ◽  
Gastric Cancer Cell ◽  
Single Chain ◽  
Single Chain Antibody ◽  
Ags Cells ◽  
Thiazolyl Blue Tetrazolium Bromide

Background:Regenerating islet-derived gene family member 4 (Reg4), a well-investigated growth factor in the regenerative pancreas, has recently been reported to be highly associated with a majority of gastrointestinal cancers. Pathological hyper-expression or artificial over-expression of Reg4 causes acceleration of tumor growth, migration, and resistance to chemotherapeutic 5-Fluorouracil (5-FU). Until now, no method has been successfully established for eliminating the effects of Reg4 protein.Methods:This study reports the production of an engineered immunoglobin, a single-chain variable fragment (scFv-Reg4), to specifically bind Reg4 and block the bioactivity. The complementary-determining regions (CDRs) against Reg4 were assigned using MOE and ZDOCK servers. The binding affinity (KD) was determined by bio-layer interferometry (BLI). MKN45 and AGS cell proliferation was determined by Thiazolyl blue tetrazolium bromide (MTT) method and the cell apoptosis was detected by flow cytometry assay.Results:The KD of scFv-Reg4 to Reg4 was determined to be 1.91×10-8. In MKN45 and AGS cell lines, scFv- Reg4 depressed Reg4-stimulated cell proliferation and the inhibitory rates were 27.7±1.5% and 17.3±2.6%, respectively. Furthermore, scFv significantly enhanced 5-FU-induced cell death, from 23.0±1.0% to 28.4±1.2% in MKN45 and 28.2±0.7% to 36.6±0.6% in AGS cells. Treatment with scFv alone could lyse cancer cells to a certain extent, but no significance has been observed.Conclusion:The single-chain antibody (scFv-Reg4) significantly inhibited gastric cancer cell proliferation and synergistically enhanced the lethal effect of 5-FU. Thus, traditional chemo-/radio- therapeutics supplemented with scFv-Reg4 may provide advances in the strategy for gastrointestinal cancer treatment.

scholarly journals Ribosome Display Technology: Applications in Disease Diagnosis and Control

Antibodies ◽  
2020 ◽  
Vol 9 (3) ◽  
pp. 28
Author(s):  
Adinarayana Kunamneni ◽  
Christian Ogaugwu ◽  
Steven Bradfute ◽  
Ravi Durvasula
Keyword(s):  
In Vitro Selection ◽  
Low Cost ◽  
Disease Diagnosis ◽  
Single Chain ◽  
Single Chain Antibody ◽  
Ribosome Display ◽  
Technology Applications ◽  
Display Technology ◽  
And Control

Antibody ribosome display remains one of the most successful in vitro selection technologies for antibodies fifteen years after it was developed. The unique possibility of direct generation of whole proteins, particularly single-chain antibody fragments (scFvs), has facilitated the establishment of this technology as one of the foremost antibody production methods. Ribosome display has become a vital tool for efficient and low-cost production of antibodies for diagnostics due to its advantageous ability to screen large libraries and generate binders of high affinity. The remarkable flexibility of this method enables its applicability to various platforms. This review focuses on the applications of ribosome display technology in biomedical and agricultural fields in the generation of recombinant scFvs for disease diagnostics and control.

scholarly journals Llama-Derived Single-Chain Antibody Fragments Directed to Rotavirus VP6 Protein Possess Broad Neutralizing Activity In Vitro and Confer Protection against Diarrhea in Mice

2008 ◽  
Vol 82 (19) ◽  
pp. 9753-9764 ◽  
Author(s):  
Lorena Garaicoechea ◽  
Aurelien Olichon ◽  
Gisela Marcoppido ◽  
Andrés Wigdorovitz ◽  
Marina Mozgovoj ◽  
...  
Keyword(s):  
Capsid Protein ◽  
Binding Capacity ◽  
Antibody Fragments ◽  
Target Antigen ◽  
Single Chain ◽  
Single Chain Antibody ◽  
Group A Rotavirus ◽  
Group A

ABSTRACT Group A rotavirus is one of the most common causes of severe diarrhea in human infants and newborn animals. Rotavirus virions are triple-layered particles. The outer capsid proteins VP4 and VP7 are highly variable and represent the major neutralizing antigens. The inner capsid protein VP6 is conserved among group A rotaviruses, is highly immunogenic, and is the target antigen of most immunodiagnosis tests. Llama-derived single-chain antibody fragments (VHH) are the smallest molecules with antigen-binding capacity and can therefore be expected to have properties different from conventional antibodies. In this study a library containing the VHH genes of a llama immunized with recombinant inner capsid protein VP6 was generated. Binders directed to VP6, in its native conformation within the viral particle, were selected and characterized. Four selected VHH directed to conformational epitopes of VP6 recognized all human and animal rotavirus strains tested and could be engineered for their use in immunodiagnostic tests for group A rotavirus detection. Three of the four VHH neutralized rotavirus in vivo independently of the strain serotype. Furthermore, this result was confirmed by in vivo partial protection against rotavirus challenge in a neonatal mouse model. The present study demonstrates for the first time a broad neutralization activity of VP6 specific VHH in vitro and in vivo. Neutralizing VHH directed to VP6 promise to become an essential tool for the prevention and treatment of rotavirus diarrhea.

scholarly journals Multimeric humanized varicella-zoster virus antibody fragments to gH neutralize virus while monomeric fragments do not

2001 ◽  
Vol 82 (8) ◽  
pp. 1959-1963 ◽  
Author(s):  
P. D. Drew ◽  
M. T. Moss ◽  
T. J. Pasieka ◽  
C. Grose ◽  
W. J. Harris ◽  
...  
Keyword(s):  
Varicella Zoster Virus ◽  
Cultured Cells ◽  
Cross Reactivity ◽  
Antibody Fragments ◽  
Single Chain ◽  
Single Chain Antibody ◽  
Neutralizing Activity ◽  
Varicella Zoster ◽  
Complementarity Determining Region

Murine monoclonal antibody 206 (MAb mu206) binds to gH, the varicella-zoster virus (VZV) fusogen, neutralizing the virus in vitro in the absence of complement and inhibiting cell-to-cell spread and egress of VZV in cultured cells. We have humanized this antibody to generate MAb hu206 by complementarity determining region grafting. MAb hu206 retained binding and in vitro neutralizing activity, as well as cross-reactivity with ten different VZV strains. Single-chain antibody fragments (scAb) derived from MAb hu206 were produced in Escherichia coli. These scAb retained the binding properties of the whole antibody. However, monomeric scAb exhibited markedly reduced neutralizing activity compared to the bivalent parental MAb hu206. Shortening the peptide linker joining the VH to the Vκ domain from 14 to 5 or even 0 residues encouraged multimerization and increased neutralizing efficacy. The fact that Fab fragments enzymatically generated from whole MAb hu206 lost their neutralizing potency lent support to the proposal that valency is important for VZV neutralization at this epitope.

scholarly journals Antibodies that potently inhibit or enhance SARS-CoV-2 spike protein-ACE2 interaction isolated from synthetic single-chain antibody libraries

2020 ◽  
Author(s):  
Matthew D. Beasley ◽  
Sanja Aracic ◽  
Fiona M. Gracey ◽  
Ruban Kannan ◽  
Avisa Masarati ◽  
...  
Keyword(s):  
Vaccine Development ◽  
Spike Protein ◽  
Receptor Binding Domain ◽  
Single Chain ◽  
Single Chain Antibody ◽  
High Affinity ◽  
Scfv Library ◽  
Antibody Libraries ◽  
Prophylactic Use

AbstractAntibodies with high affinity against the receptor binding domain (RBD) of the SARS-CoV-2 S1 ectodomain were identified from screens using the Retained Display™ (ReD) platform employing a 1 × 1011 clone single-chain antibody (scFv) library. Numerous unique scFv clones capable of inhibiting binding of the viral S1 ectodomain to the ACE2 receptor in vitro were characterized. To maximize avidity, selected clones were reformatted as bivalent diabodies and monoclonal antibodies (mAb). The highest affinity mAb completely neutralized live SARS-CoV-2 virus in cell culture for four days at a concentration of 6.7 nM, suggesting potential therapeutic and/or prophylactic use. Furthermore, scFvs were identified that greatly increased the interaction of the viral S1 trimer with the ACE2 receptor, with potential implications for vaccine development.

A Novel Anti CD23 Fully Human Monoclonal Antibody Potentially Useful for B-CLL Therapy.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 5047-5047
Author(s):  
Marc Delcommenne ◽  
Hans-Georg Klingemann ◽  
Stephanie A. Gregory
Keyword(s):  
Monoclonal Antibody ◽  
B Lymphocytes ◽  
Human Monoclonal Antibody ◽  
Myeloma Cell ◽  
Lymphocytic Leukemia ◽  
Single Chain ◽  
Single Chain Antibody ◽  
Normal Peripheral Blood ◽  
Human Igg1

Abstract B-cell chronic lymphocytic leukemia (B-CLL) is one of the most common hematological malignancies and is, in most cases, characterized by an increased expression of CD23 on the cell surface. Since cross-linking CD23 induces B-CLL apoptosis, it is an attractive target for B-CLL antibody-based immunotherapy. In this study we show that an anti-CD23 human IgG1 monoclonal antibody, C6F5, may be useful in treating B-CLL. This antibody is derived from the human single chain antibody (scFv) C6F5 that was originally raised against the RPMI-8226 multiple myeloma cell line using the antibody phage display technique. While the C6F5 scFv did not bind to other myeloma cell lines, it was able to bind weakly to normal peripheral blood B lymphocytes and strongly to EBV transformed B cells and B-CLL cells. The antigen recognized by C6F5 was also upregulated on B lymphocytes that had been stimulated by CD40 ligand. Immunoprecipitations by the scFv C6F5 identified a protein of 45 kDa which co-migrated with CD23. Furthermore, this protein was recognized by an anti-CD23 mouse mAb in Western blot analyses. Immunofluorescence staining with the C6F5 scFv was inhibited if cells were preincubated with an anti-CD23 polyclonal antiserum. Taken together, these results verify that C6F5 recognizes CD23. The VH and VL regions of C6F5 antibody were then cloned into a baculovirus transfer vector encoding the human IgG1 heavy and light chains so that fully human C6F5 IgG1 antibody could be produced in baculovirus infected SF9 cells. Since C6F5 binding specificity was preserved in the IgG1 format, this antibody is ready to be tested in in vitro cytotoxic assays against B-CLL cells.

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