中国兽医杂志2025，Vol.61Issue(3)：22-28,7.DOI:10.20157/j.cnki.zgsyzz.2025.03.003
含分子内佐剂的ASFV重组蛋白制备和免疫效果评价
Preparation and Immune Effect Evaluation of ASFV Recombinant Protein Containing Intramolecular Adjuvant
摘要
Abstract
This study aimed to explore the effect of the combined action of the intramolecular adjuvants Tuftsin and tetanus toxin peptide TT on humoral and cellular immunity induced by recombinant African swine fever virus(ASFV)protein P753.Using bioinformatics software,dominant epitopes of ASFV structural proteins P72,P54,and P30 were identified,and recombinant vectors pET-28a(+)-p753 and pET-28a(+)-p753-Tuftsin-TT were constructed.The recombinant proteins were expressed using an Escherichia coli expression system,purified,and renatured.Immunogenicity and immune reactivity of the recombinant proteins were verified by Western blot and Dot-enzyme-linked immunosorbent assay(Dot-ELISA).Mice were immunized with the two recombinant proteins mixed with the commercial veterinary adjuvant ISA 61 VG.The humoral and cellular immune responses were evaluated by detecting serum antibody levels,spleen lymphocyte proliferation,and cytokine(IL-2,IL-4,and IFN-γ)secretion levels.The results showed that both recombinant proteins were expressed as inclusion bodies under conditions of 37℃and 0.5 mmol/L IPTG induction for 4 hours,with the highest expression levels.After purification and renaturation,both proteins displayed immunogenicity and immune reactivity.After immunization,the antibody levels in the P753 group and P753-Tuftsin-TT group reached a plateau at 42 days post-primary immunization,with both groups showing higher antibody levels than the PBS group.Among them,the P753-Tuftsin-TT group exhibited the highest antibody levels,and the antigen stimulation index and cytokine(IL-2,IL-4,and IFN-γ)levels were significantly higher than those in the P753 group(P<0.000 1).The findings suggest that the combined use of intramolecular adjuvants Tuftsin and TT significantly enhances the humoral and cellular immune responses induced by P753 recombinant antigen,providing a reference for the development of ASFV subunit vaccine adjuvants.关键词
非洲猪瘟病毒(ASFV)/分子内佐剂/亚单位疫苗/体液免疫/细胞免疫Key words
African swine fever virus(ASFV)/intramolecular adjuvant/subunit vaccine/humoral immunity/cellular immu-nity分类
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李玉林,周方方,闫生辉,于巧玲,王继创,石盼盼,王云龙..含分子内佐剂的ASFV重组蛋白制备和免疫效果评价[J].中国兽医杂志,2025,61(3):22-28,7.基金项目
河南省重点研发与推广专项(222102110159) （222102110159）
