Design and construction of immune phage antibody library against Tetanus neurotoxin: Production of single chain antibody fragments.

Publisher: IOS Press Country of Publication: Netherlands NLM ID: 9711270 Publication Model: Print Cited Medium: Internet ISSN: 1875-869X (Electronic) Linking ISSN: 10932607 NLM ISO Abbreviation: Hum Antibodies Subsets: MEDLINE

Publication: Amsterdam : IOS Press
Original Publication: Wilton, CT : Forefront Pub., c1997-

Peptide Library*; Antibodies, Bacterial/*biosynthesis ; Antibodies, Monoclonal/*biosynthesis ; Immunoglobulin G/*biosynthesis ; Single-Chain Antibodies/*biosynthesis ; Tetanus/*prevention & control; Antibodies, Bacterial/isolation & purification ; Antibodies, Bacterial/pharmacology ; Antibodies, Monoclonal/isolation & purification ; Antibodies, Monoclonal/pharmacology ; B-Lymphocytes/immunology ; B-Lymphocytes/microbiology ; Clostridium tetani/immunology ; Clostridium tetani/pathogenicity ; Drug Design ; Enzyme-Linked Immunosorbent Assay ; Humans ; Immunoglobulin G/isolation & purification ; Immunoglobulin G/pharmacology ; Metalloendopeptidases/antagonists & inhibitors ; Metalloendopeptidases/chemistry ; Metalloendopeptidases/immunology ; Protein Binding ; Protein Engineering ; Single-Chain Antibodies/isolation & purification ; Single-Chain Antibodies/pharmacology ; Tetanus/immunology ; Tetanus/microbiology ; Tetanus Toxin/antagonists & inhibitors ; Tetanus Toxin/chemistry ; Tetanus Toxin/immunology ; Tetanus Toxoid/antagonists & inhibitors ; Tetanus Toxoid/chemistry ; Tetanus Toxoid/immunology

Background: Tetanus neurotoxin (TeNT) is composed of a light (LC) and heavy chain (HC) polypeptides, released by anaerobic bacterium Clostridium tetani and can cause fatal life-threatening infectious disease. Toxin HC and LC modules represents receptor binding and zinc metalloprotease activity, respectively. The passive administration of animal-derived antibodies against tetanus toxin has been considered as the mainstay therapy for years. However, this treatment is associated with several adverse effects due to the presence of anti-isotype antibodies.
Objective: In the present study, we have produced the fully human single chain antibody fragments (HuScFv) from two human antibody phage display libraries.
Material and Methods: Twenty-four different HuscFvs were isolated from two anti TeNT immune libraries. Our produced human ScFv (HuScFv) were converted to IgG platform and analyzed regarding their specific reactivity to TeNT.
Results: All of the selected scFvs have the same VL but different VH. Three HuscFvs from the first library (TTX15, 51, 75) and two HuscFvs from the second library (TTX16, 20) were chosen to convert to IgG1 using pOptiVEC and pcDNA3.3 systems. Production of IgG1 from transfected DG44 and binding capacity of them to tetanus toxin and toxoid were measured by ELISA. ELISA results showed no detectable production of TTX16 and TTX20 IgG1. Although, TTX51 and TTX75 were converted and produced as IgG1, no reactivity to tetanus toxin and toxoid was observed. However, TTX15 was successfully produced as whole IgG1 platform with reactivity to both tetanus toxin and toxoid. The latter would be an appropriate replacement for conventional polyclonal antibodies if would meet the further characterization including specificity determination, affinity measurement and toxin neutralizing assays.
Conclusion: Our results demonstrated production of functional IgG1 derived from TTX15 scFv and might be an appropriate replacement for polyclonal Tetabulin but it needs further characterization.

Keywords: ScFv; Tetanus neurotoxin; neutralizing antibodies

0 (Antibodies, Bacterial)
0 (Antibodies, Monoclonal)
0 (Immunoglobulin G)
0 (Peptide Library)
0 (Single-Chain Antibodies)
0 (Tetanus Toxin)
0 (Tetanus Toxoid)
11032-48-7 (tetanospasmin)
EC 3.4.24.- (Metalloendopeptidases)

Date Created: 20160730 Date Completed: 20170310 Latest Revision: 20170310

20250114

10.3233/HAB-150287

27472865