Biotechnology letters · 2016
Screening, tandem expression and immune activity appraisal of Deinagkistrodon acutus (pit viper) venom mimotopes from a phage display 12-mer peptide library.
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Guo Guoning, Cao Yuliang, Zhu Guoyan, Tian Zhu, Gou Yajun, Chen Cong, Liu Minghua
Аннотация
OBJECTIVE: To design a specific polyclonal antibody against Deinagkistrodon acutus venom (DA-pAb) by immunizating New Zealand white rabbits.
RESULTS: The IgG fraction was purified by affinity chromatography, and specific antibodies were purified by immunoaffinity chromatography. Polyclonal antibodies were subjected to ELISA and western blotting to evaluate their immune reactivity. We identified the mimotopes by screening a phage display 12-mer peptide library against D. acutus venom. After three rounds of biopanning with DA-pAb, 30 positive clones were identified. Eighteen phage clones were sequenced, and their corresponding amino acid sequences were deduced. Additional immunoassays with the peptides and DA-pAb identified five sequences as possible epitopes. Recombinant antigens synthesized with the five epitopes were used for the immunization of BALB/c mice.
CONCLUSION: The antibodies induced by these peptides recognized the recombinant antigen and D. acutus venom and protected mice against the hemorrhagic effects of the venom.
RESULTS: The IgG fraction was purified by affinity chromatography, and specific antibodies were purified by immunoaffinity chromatography. Polyclonal antibodies were subjected to ELISA and western blotting to evaluate their immune reactivity. We identified the mimotopes by screening a phage display 12-mer peptide library against D. acutus venom. After three rounds of biopanning with DA-pAb, 30 positive clones were identified. Eighteen phage clones were sequenced, and their corresponding amino acid sequences were deduced. Additional immunoassays with the peptides and DA-pAb identified five sequences as possible epitopes. Recombinant antigens synthesized with the five epitopes were used for the immunization of BALB/c mice.
CONCLUSION: The antibodies induced by these peptides recognized the recombinant antigen and D. acutus venom and protected mice against the hemorrhagic effects of the venom.