2014•PubMedRequires access
[Preparation and application of polyclonal antibody against chicken high mobility group box 1 protein].
Na Zhang, Kun Qian, Mingyue Zhu, Ai-jun Gao, Hongxia Shao, Wenjie Jin, Aijian Qin
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Abstract
OBJECTIVE: To prepare an affinity purified polyclonal antibody against chicken high mobility group box 1 protein (chHMGB1) and put it in a practical application. METHODS: The full-length chHMGB1 cDNA was amplified by reverse transcription PCR (RT-PCR). The specific peptides was selected by sequence alignment and conjugated with KLH. Then it was used to immunize the New Zealand rabbits to obtain the anti-serum. The polyclonal antibody was purified by protein A column and peptide-conjugated Seprose4B sequentially. ELISA, Western blotting, indirect fluorescence assay (IFA) and immunohistochemistry were used for the identification and application of the affinity purified polyclonal antibody. RESULTS: These examinations demonstrated the successful generation of the affinity purified polyclonal antibody against chHMGB1 protein and the specificity of the polyclonal antibody to both the recombinant protein and the natural chHMGB1 protein. CONCLUSION: The affinity purified polyclonal antibody was successfully prepared.