| Sulpholipopolysaccharides(SLP)is the product of esterified and sulfated polysaccharide.Compared to the pure polysaccharide,ester-chain gene and sulphoacid gene are added to its molecular structure.Therefore,it's both lipophilic and hydrophilic,and can be used as a surfactant.Surveys abroad show that adjuvant activity of the sulfated ficoll is related to esterification and sulfation.The SLP with low sulfation degree and high esterification degree has better adjuvant activity.As the surfactant,SLP plays as the role of emulsification during the process of the production of oil-in-water adjuvant.Research shows that the adjuvant formed after SLP emulsificate squalene oil-in-water adjuvant has better immune activity than normal oil-in-water adjuvant.In this study,through orthogonal screening,SLP Ficoll and SLP Cyclodextrins with low sulfation and high esterification could be produced by Ficoll 400 and ?-Cyclodextrins.Then the oil-in-water adjuvants were produced by them.Through the interval test in mice and swine,The effect on animal lymphocyte proliferation,cytokines,antibody and its subtype of SLP Ficoll and SLP Cyclodextrins were studied through the interval test in mice and swine.The aim of the study was to develop the new,efficient and low toxic oil-in-water adjuvant to replace the conventional oil adjuvant.The test including four parts as following:Experiment I Optimization on preparation conditions of Sulpholipo-Ficoll and Sulpholipo-CyclodextrinsSulpholipo-Ficoll and Sulpholipo-Cyclodextrins were prepared by lipid and sulphate groups coupled to polysaccharide.To optimize the preparation conditions of SLP,the influence of lipid per monosaccharide unit and sulphate groups per monosaccharide unit on the number of lipid and sulphate groups per monosaccharide was studied by orthogonal screening.The data was analyzed with Orthogonality Experiment Assistant,the optimum preparation conditions were as follows:lipid per monosaccharide unit was 3.6,sulphate groups per monosaccharide unit was 1.0,temperature was 60 ?.The results showed that Sulpholipo-Ficoll was prepared as the optimal preparation condition with the number of lipid and sulphate groups per monosaccharide were 3.24 and 0.40.Sulpholipo-Cyclodextrins was prepared as the optimal preparation condition with the number of lipid and sulphate groups per monosaccharide were 1.70 and 1.32.Experiment ? Studies on adjuvanticity of OVA immunized mices of two Oil-in-water adjuvantsIn order to research the adjuvanticity of two.oil-in-water adjuvants on OVA immunized mices,randomly dividing 150 5-week mices into 6 groups,and giving subcutaneous immunization with OVA taking two sulpholipopolysaccharides as adjuvants.Then the mice were boosted after 6 weeks.Blood samples were collected through taking away eyeballs after 7-day,14-day,21-day and 28-day since the first immunization.Then the concentration changes of IgG,IL-4 and IFN-? through ELISA were determined.Six mice were killed in each group and the splenic lymphocytes were separated.The lymphocytes proliferation of T cell and B cell were determined through MTT.The result showed that in some time,the SLP oil-in-water adjuvant could prominently increase the concentration of IgG and IL-2,and improve the immune activity of antigen.It has good prospect to develop and apply as new oil-in-water adjuvant.Experiment ? Studies on adjuvanticity of Porcine circovirus ?immunized mices of two Oil-in-water adjuvantsIn order to research the adjuvanticity of two oil-in-water adjuvants on Porcine Circovirus ? immunized mices,the50 5-week Balb/c female mices were randomly divided into 5 groups,and given subcutaneous immunization with Porcine Circovi ? vaccine,taking two sulpholipopolysaccharides as adjuvants.The blood samples were collectedfrom orbital vein after 7-day,14-day,21-day,28-day and 35-day since the immunization.Then the concentration changes of typical Porcine Circovi ? IgG,antibody subtype IgG1 and IgG2a were tesed through ELISA.The result showed that in some time,the SLP oil-in-water adjuvant could prominently increase the concentration of typical Porcine Circovi ? IgG,antibody subtype IgG1 and IgG2a,or similar to the activity of the oil adjuvant.Overall,SLP oil-in-water adjuvant could effectively improve the immune activity of antigen and had good prospect to develop and apply as new oil-in-water adjuvant.Experiment ? Studies on adjuvanticity of Porcine circovirus ?immunized swine of two Oil-in-water adjuvantsIn order to research the adjuvanticity of two oil-in-water adjuvants on Porcine Circovirus ? immunized swine,20 30-day swine were randomly divided into 4 groups,and giving jugular subcutaneous immunization with Porcine Circovi ? vaccine taking two sulpholipopolysaccharides as adjuvants.Collecting blood samples from Orbital vein after 28-day since the immunization.Then the concentration of typical Porcine Circovi ? IgG were test through ELISA.The result showed that in some time,the SLP oil-in-water adjuvant could prominently increase the concentration of typical Porcine Circovi ? IgG,which was similar to the activity of Ringpu contrasted vaccine.The results indicated that SLP oil-in-water adjuvant couldeffectively improve the immune activity of the antigen,and would provide the development and application of new oil-in-water adjuvant with research basis. |
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