The aim of this study is to explore the immunogenicity of African swine fever virus p37 recombinant protein in mice.C57BL/6J mice were immunized subcutaneously in the abdomen using p37 recombinant protein as antigen.The second immunization was performed 21 d after the first immunization.Serum-specific antibody levels were detected by ELISA;serum cytokine levels were detected using a multifactor assay technique;mice splenic lymphocytes were isolated 7 d after sec-ondary immunization,and the number of splenic lymphocytes secreting IFN-γ after recombinant protein stimulation was detected by ELISpot;and the ratio of CD4+T cells to CD8+T cells was detected by flow cytometry.The results of indirect ELISA showed that p37 recombinant protein could stimulate mice to produce high levels of specific antibodies;ELISpot showed that p37 recom-binant protein could significantly stimulate splenic lymphocytes to produce IFN-γ(P＜0.001)and activate cellular immune responses;the results of flow cytometry showed that it could signifi-cantly stimulate the differentiation of T-lymphocytes to CD4+T-lymphocytes(P＜0.001).In ad-dition,serum levels of IL-2,IL-4,IFN-γ,and TNF-α immune-related cytokines were significantly higher after the second immunization.Immunization of mice with p37 recombinant protein induced strong humoral and cellular immune responses with good immunogenicity,providing reference for the subsequent epitope identification and functional study of p37 protein and the antigen screening of ASF mRNA vaccine.

To explore the interaction between ASFV capsid protein M1249L and host from the host cellular perspective,M1249L was selected for constructing the bait plasmid(pGBKT7-M1249L)to screen the bone marrow-derived macrophages(BMDMs)cDNA library.After again co-transform and sequence alignment,20 candidate interacting host proteins were screened,such as IL-1β,CTSB and DNAJA3.And then,co-immunoprecipitation assay was performed to verify the interaction be-tween M1249L and host proteins.GO ontology(GO)and KEGG pathway enrichment analyses re-vealed that biological regulation,cellular communication and response to stimulus and others were enriched in biological processes.And these host proteins could share some pathways,including toll-like receptor signaling pathway and Nod-like receptor signaling pathway.Therefore,the results provides the theoretical basis for further research on the mechanism of ASFV M1249L in viral in-fection and immune regulation.

The aim of this study is to explore the immunogenicity of African swine fever virus p37 recombinant protein in mice.C57BL/6J mice were immunized subcutaneously in the abdomen using p37 recombinant protein as antigen.The second immunization was performed 21 d after the first immunization.Serum-specific antibody levels were detected by ELISA;serum cytokine levels were detected using a multifactor assay technique;mice splenic lymphocytes were isolated 7 d after sec-ondary immunization,and the number of splenic lymphocytes secreting IFN-γ after recombinant protein stimulation was detected by ELISpot;and the ratio of CD4+T cells to CD8+T cells was detected by flow cytometry.The results of indirect ELISA showed that p37 recombinant protein could stimulate mice to produce high levels of specific antibodies;ELISpot showed that p37 recom-binant protein could significantly stimulate splenic lymphocytes to produce IFN-γ(P＜0.001)and activate cellular immune responses;the results of flow cytometry showed that it could signifi-cantly stimulate the differentiation of T-lymphocytes to CD4+T-lymphocytes(P＜0.001).In ad-dition,serum levels of IL-2,IL-4,IFN-γ,and TNF-α immune-related cytokines were significantly higher after the second immunization.Immunization of mice with p37 recombinant protein induced strong humoral and cellular immune responses with good immunogenicity,providing reference for the subsequent epitope identification and functional study of p37 protein and the antigen screening of ASF mRNA vaccine.

To explore the interaction between ASFV capsid protein M1249L and host from the host cellular perspective,M1249L was selected for constructing the bait plasmid(pGBKT7-M1249L)to screen the bone marrow-derived macrophages(BMDMs)cDNA library.After again co-transform and sequence alignment,20 candidate interacting host proteins were screened,such as IL-1β,CTSB and DNAJA3.And then,co-immunoprecipitation assay was performed to verify the interaction be-tween M1249L and host proteins.GO ontology(GO)and KEGG pathway enrichment analyses re-vealed that biological regulation,cellular communication and response to stimulus and others were enriched in biological processes.And these host proteins could share some pathways,including toll-like receptor signaling pathway and Nod-like receptor signaling pathway.Therefore,the results provides the theoretical basis for further research on the mechanism of ASFV M1249L in viral in-fection and immune regulation.

African swine fever(ASF)is a highly contagious disease caused by the African swine fe-ver virus(ASFV).To evaluate the immunogenicity of the pp62(CP530R)protein of ASFV,the recombinant CP530R protein was expressed in HEK 293F cells transfected with the plasmid pMAL-Fc-CP530R.Six-week-old female C57BL/6J mice were immunized with 10 μg of purified pp62 protein via subcutaneous injection,followed by a booster immunization with the same dosage at 21 days(enhanced).The humoral and cellular immune responses in the mice were then assessed using ELISA,flow cytometry,and ELISpot assays.Western blot analysis confirmed that the pp62 protein was successfully expressed,with a molecular weight of approximately 118.5 kDa.ELISA results indicated that a high level of specific antibodies was detected in the immunized mice,with antibody titers reaching up to 1∶1 638 400 at 7 days after the secondary immunization.The pro-portion of CD8+T lymphocytes in the immunized mice increased compared to the control group(P＜0.05).Results from the Q-PlexTM Mouse Cytokine Screen demonstrated that the secretion levels of IFN-γ,IL-2,IL-4,and IL-10 in serum were significantly upregulated in the immunized mice following secondary immunization(P＜0.001).In summary,these findings indicate that the pp62 protein can significantly stimulate both humoral and cellular immunity in mice,laying the groundwork for further studies on the function of the ASFV pp62 protein and the identification of novel vaccine antigens for ASF.

African swine fever(ASF)is a highly contagious disease caused by the African swine fe-ver virus(ASFV).To evaluate the immunogenicity of the pp62(CP530R)protein of ASFV,the recombinant CP530R protein was expressed in HEK 293F cells transfected with the plasmid pMAL-Fc-CP530R.Six-week-old female C57BL/6J mice were immunized with 10 μg of purified pp62 protein via subcutaneous injection,followed by a booster immunization with the same dosage at 21 days(enhanced).The humoral and cellular immune responses in the mice were then assessed using ELISA,flow cytometry,and ELISpot assays.Western blot analysis confirmed that the pp62 protein was successfully expressed,with a molecular weight of approximately 118.5 kDa.ELISA results indicated that a high level of specific antibodies was detected in the immunized mice,with antibody titers reaching up to 1∶1 638 400 at 7 days after the secondary immunization.The pro-portion of CD8+T lymphocytes in the immunized mice increased compared to the control group(P＜0.05).Results from the Q-PlexTM Mouse Cytokine Screen demonstrated that the secretion levels of IFN-γ,IL-2,IL-4,and IL-10 in serum were significantly upregulated in the immunized mice following secondary immunization(P＜0.001).In summary,these findings indicate that the pp62 protein can significantly stimulate both humoral and cellular immunity in mice,laying the groundwork for further studies on the function of the ASFV pp62 protein and the identification of novel vaccine antigens for ASF.