Optimization of immune scheme for SARS-CoV-2 RBD recombinant protein vaccine based on P.pastoris and the effect of different adjuvants on neutralizating antibody titer / 国际检验医学杂志

ABSTRACT

Objective To optimize the immune scheme of SARS-CoV-2 RBD recombinant protein vaccine based on P.pastoris,and investigate the effect of different adjuvants on neutralizating antibody(NAb)titer,in order to provide reference for the continuous optimization research of SARS-CoV-2 vaccine.Methods The RBD protein was selected and the corresponding gene fragment was synthesized,which was constructed into the pPICZαA plasmid,and the plasmid was integrated into the genome of P.pastoris after linear transforma-tion for recombinant expression.The obtained recombinant protein vaccine was combined with different adju-vants to immunize mice to evaluate its immunogenicity.Results Both the target proteins wtRBD and Delta RBD were able to achieve satisfactory overexpression through the P.pastoris system.Compared with the 42 d interval,the IgG antibody titer at the 28 d interval increased by 1.8 times(44 923 vs.80 507).After 3 doses of immunization at an interval of 28 d,the geometric mean titer of NAb for Delta variant was 2.5 times higher than that at an interval of 42 days(2 191 vs.891).After immunization with Delta RBD recombinant protein vaccine combined with aluminum adjuvant,the NAb geometric mean titer for Delta variants reached 32 255(2 167-88 084).When using 5 μg or 30 μg Delta RBD immunization,the NAb titers of the aluminum adju-vant+CpG adjuvant group were about 10 times higher than those of the aluminum adjuvant group alone.Af-ter the third immunization,there was no significant difference in Delta RBD specific IgG titers between the 5 μg antigen group and the 30 μg antigen group(P＞0.05).Conclusion Both wtRBD and Delta RBD prepared based on P.pastoris could be used as effective antigens,with three doses of vaccine administered at a 28 day in-terval being the most effective.The combined immunization of Delta RBD recombinant protein with aluminum adjuvant+CpG adjuvant could obtain higher titers of NAb to exert immune effects on SARS-CoV-2 and its va-riants,providing some reference for the continuous optimization research of SARS-CoV-2 vaccines.