Background: The emergence of infectious agents enforces rapid prophylactic/therapeutic development. Accordingly, multi-epitope vaccines represent a milestone in the second-generation vaccines to break the SARS-CoV-2 transmission chain, a zoonotic virus, and decrease long coronavirus (COVID-19) complications. Thus, this study aimed to present a potent anti-SARS-CoV-2 pan-species vaccine candidate via computational workflow to address this hypothesis: Could a multi-epitope construct provoke effective anti-SARS-CoV-2 protection as a bioinformatics solution? Methods: The B-cell and T-cell epitopes of the S1 domain, N, Nsp4, Nsp12, and ORF6 proteins were predicted regarding the significant role of viral proteins in the infection cycle. Then, the final construct was designed as (1) scaffolding by the top-ranked epitopes, β-defensin, His6-tag, (2) structural modeling/refinement, (3) molecular docking/dynamics simulation, and (4) validation. Results: The 15 linear B-cell, 7 helper T-cell, and 11 cytotoxic T-cell epitopes were embedded within the construct, all located outside the hyper-variable regions of target proteins. Following the quality assessment, its best binding affinity was observed with TLR4, TLR8, HLA-DRB1*01:01, HLA-A*03:01, and ACE2. The strong interactions were also observed with a validated single-chain variable fragment as a preliminary in silico indication about its possible in vivo efficiency. Moreover, the messenger RNA stability and robust immune stimulation were verified. Not only was non-significant homology found between the multi-epitope vaccine construct and some viral hosts/reservoir proteomes, but also significant coverage was detected between human immune receptors and their counterparts in domestic, wildlife, and captive animals. Therefore, it could be a sign of the possible efficiency of the construct as a pan-species vaccine. Conclusion: Despite the end of the COVID-19 pandemic, a combinatorial workflow was introduced to design a vaccine with potential application in various species, whereas many other studies have not employed all of these criteria, and according to our information, this is one of the few works to introduce such constructs.

Purpose: Among all known human coronaviruses, some viruses (e.g., SARS-CoV-2) cause severe pneumonia or even death. With the regard to its spread and the importance of its rapid identification/treatment, and because pAbs are relatively cheap, able to bind to more sites on antigens and even neutralize them, this study was done for the production and purification of anti-SARS-CoV-2 polyclonal antibodies (pAb) in rats. Methods: Viral antigen purification was performed by PEG/NaCl precipitation. The efficiency of the sucrose cushion method was also investigated to produce a purer antigen. Immunization was done and antibody purification was performed by ammonium sulfate precipitation (33%), dialysis, and ion-exchange chromatography. Western blotting and enzyme-linked immunosorbent assay (ELISA) were performed to verify the antibody specificity. All data were analyzed by SPSS software. Results: The results showed that the amount of concentrated virus increased with the increase of PEG concentration. Moreover, the sucrose cushion method increased its purity. Besides, induction of immune response in rats for pAb production with high titers was reached via these antigens and ELISA/western blot results indicated a suitable antibody-antigen interaction. Additionally, it was shown that ion-exchange chromatography could be a suitable technique for IgG purification. Conclusion: Herein, we presented a simple and cheap method for the purification of whole viral particles with relatively high quality. The results verified that these antigens could elicit a good immune response in the rat. The obtained pAbs showed a good specificity toward SARS-CoV-2 antigens. Accordingly, this study proposes a promising method for viral vaccine development.