Production and Characterization of Neutralizing Antibodies against Bungarus multicinctus Snake Venom.
The venom of the banded krait (Bungarus multicinctus), one of the major venomous species in Taiwan, contains neurotoxic venom proteins (B. multicinctusproteins) that pose a serious medical problem in tropical and subtropical countries. Even though horse-derived serum is an efficient therapy against snake venom, it is associated with a high cost and side effects. Therefore, developing a more cost-effective alternative treatment option is highly envisaged. In this study, chickens were immunized withB. multicinctusproteins, and polyclonal immunoglobulin Y (IgY) antibodies were purified from eggs. IgY showed a binding activity toB. multicinctusproteins that was similar to horse antivenin, and its titer in chickens lasted for at least 6 months. We constructed two antibody libraries by phage display antibody technology, which contain 1.0 × 107and 2.9 × 108transformants, respectively. After biopanning, a phage-based enzyme-linked immunosorbent assay (ELISA) indicated that 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 toB. multicinctusproteins but not to the venomous proteins of other snakes. Most importantly, polyclonal IgY demonstrated a similar neutralization efficiency as did horse-derived antivenin in mice that were injected with a minimum lethal dosage (MLD) of venom proteins. A mixture of several monoclonal anti-B. multicinctusscFv antibodies was also able to partially inhibit the lethal effect on mice. We profoundly believe that IgY and scFv antibodies can be applied in developing diagnostic agents for wound exudates and as an alternative treatment for snakebite envenomation in the future.IMPORTANCESnake envenomation is one of the global medical issues of 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. A mixture of scFv antibodies was able to partially inhibit the lethal effect in mice that were injected with lethal dosages of venom proteins and prolong their survival time. We believe that chicken-derived IgY and scFv antibodies have great potential for the development of diagnostic agents for wound exudates and therapeutic agents against snake envenomation in the future.
Graduate Institute of Medical Sciences, College of Medicine, Taipei Medical University, Taipei, Taiwan.
Chi-Hsin
http://orcid.org/0000-0002-3412-7400
CH
Lee
The Center of Translational Medicine, Taipei Medical University, Taipei, Taiwan.|Ph.D. 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.
Yu-Ching
YC
Lee
Graduate 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.
Sy-Jye
SJ
Leu
Graduate 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.
Liang-Tzung
LT
Lin
Center for Research, Diagnostics and Vaccine Development, Centers for Disease Control, Ministry of Health and Welfare, Taipei, Taiwan.
Jen-Ron
JR
Chiang
Section of Molecular Biology, Department of Medical Techniques, Taipei City Hospital, Renai Branch, Taipei, Taiwan.
Wei-Jane
WJ
Hsu
Core Facility for Antibody Generation and Research, Taipei Medical University, Taipei, Taiwan yangyuan@tmu.edu.tw.|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.
Yi-Yuan
YY
Yang
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