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Production and Characterization of Neutralizing Antibodies against Bungarus multicinctus Snake Venom.

Posted by on in 2016
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Lee CH1Lee YC2Leu SJ3Lin LT3Chiang JR4Hsu WJ5Yang YY6. 2016. Appl Environ Microbiol. . pii: AEM.01876-16. [Epub ahead of print]

 

1Graduate Institute of Medical Sciences, College of Medicine, Taipei Medical University, Taipei, Taiwan.
2The Center of Translational Medicine, Taipei Medical University, Taipei, Taiwan. PhD Program for Biotechnology in Medicine, College of Medical Science and Technology, Taipei Medical University, Taipei, Taiwan. Core Facility for Antibody Generation and Research, Taipei Medical University, Taipei, Taiwan.
3Graduate Institute of Medical Sciences, College of Medicine, Taipei Medical University, Taipei, Taiwan. Department of Microbiology and Immunology, School of Medicine, College of Medicine, Taipei Medical University, Taipei, Taiwan.
4Center for Research, Diagnostics and Vaccine Development, Centers for Disease Control, Ministry of Health and Welfare, Taiwan.
5Section of Molecular Biology, Department of Medical Techniques, Taipei City Hospital, Renai Branch, Taipei, Taiwan.
6Core Facility for Antibody Generation and Research, Taipei Medical University, Taipei, Taiwan. School of Medical Laboratory Science and Biotechnology, College of Medical Science and Technology, Taipei Medical University, Taipei, Taiwan. Department of Laboratory Medicine, Wan Fang Hospital, Taipei Medical University, Taipei, Taiwan. yangyuan@tmu.edu.tw.

Abstract

Banded krait (Bungarus multicinctus; BM), one of the major envenomation species in Taiwan, contains neurotoxic venom proteins (BM proteins), that poses a serious medical problem in tropical and sub-tropical countries. Even though horse-derived serum is an efficient therapy against snake venom, it is associated with high cost and side effects. Therefore, developing a more cost-effective alternative treatment option is highly envisaged. In this study, chickens were immunized with BM proteins, and polyclonal immunoglobulin Y (IgY) antibodies were purified from eggs. IgY showed a similar binding activity to BM proteins as horse antivenin, and its titer in chickens lasted for at least 6 months. We constructed two antibody libraries by phage display antibody technology containing 1.0 ×107 and 2.9 × 108 transformants, respectively. After bio-panning, phage-based ELISA indicated specific clones were enriched. Thirty randomly selected clones expressing monoclonal single-chain variable-fragment (scFv) antibodies were classified into four groups with a short linker and two with a long linker. These selected scFv antibodies showed specific binding activities to BM proteins but not to venomous proteins of other snakes. Most importantly, polyclonal IgY demonstrated a similar neutralization efficiency as did horse-derived antivenin in mice injected with a minimum lethal dosage (MLD) of venom proteins. A mixture of several monoclonal anti-BM scFv antibodies was also able to partially inhibit the lethal effect on mice. We profoundly believe that the IgY and scFv antibodies can be applied in developing diagnostic agents for wound exudates and as an alternative treatment for snakebite envenomations in the future.

IMPORTANCE:

Snake envenomation is one of global medical issues in concern. Horse-derived antivenin is an effective way to treat snakebites but it is costly and occasionally causes severe side effects. In this study, we first generated and characterized IgY antibodies with neutralization activity in chickens. Subsequently, we generated a panel of monoclonal scFv antibodies using phage display antibody technology. Mixture of scFv antibodies was able to partially inhibit the lethal effect in mice injected with lethal dosage of venom proteins and prolong their survival time. We believe the chicken-derived IgY and scFv antibodies have great potential to develop diagnostic agents for wound exudates and therapeutic agents against snake envenomation in the future.

PMID:
 
27663029
 
PMCID:
 
PMC5103096
 [Available on 2017-05-09]
 
DOI:
 
10.1128/AEM.01876-16
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