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Preparation, Purification and Identification of Bacteriophage Qβ Virus-like Particles |
CHEN Xiu-yue,ZHOU Wen-feng,HE Qing,SU Bing,ZOU Ya-wen() |
College of Veterinary Medicine, Hunan Agricultural University, Hunan Provincial Key Laboratory of Protein Engineering in Animal Vaccines, Research & Development Center for Animal Reverse Vaccinology of Hunan Province, Changsha 410128, China |
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Abstract Objective: The self-assembly ability of phage Qβ virus-like particles (VLPs),prepared by Escherichia coli (E. coli) prokaryotic expression system was verified. To determine their immunogenicity, New Zealand rabbits were immunized with purified Qβ VLPs, and them ability to enter mammalian cells was also detected.Methods: The pET-28-Qβ-CP plasmid was constructed, and Qβ VLPs were produced by E. coli expression system. Qβ VLPs were purified by sucrose density gradient centrifugation and gel filtration (Sephacryl S-400 column). The particle morphology of Qβ VLPs were observed by transmission electron microscopy (TEM). Antisera were obtained from the rabbits immunized with the VLPs, along with or without adjuvant, and subsequently, antibodies were purified by Protein G resin. The specificity was determined by Western blot. The indirect immunofluorescence assay (IFA) was used to detect the entry of Qβ VLPs into cells.Results: High purity Qβ VLPs were obtained. TEM results showed that substantial VLPs were observed with diameter of about 28 nm. Western blot showed that Qβ VLPs were specifically recognized by anti-rabbit polyclonal antibodies. Adjuvant had no negative effect on the production of antibody. Based on IFA, the results showed that Qβ VLPs entered various mammalian cells.Conclusion: Qβ VLPs were successfully produced in this study, and they would have great potential as carriers to facilitate the development of future vaccine based on Qβ VLPs.
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Received: 15 March 2021
Published: 03 August 2021
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Corresponding Authors:
Ya-wen ZOU
E-mail: yawenzou@stu.hunau.edu.cn
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