| Nano-calcium phosphate (NCaP) not only had CaP’s characteristics such as biocompatibility, biodegradability, non-toxicity etc., also had some advantages of nanoparticles as adsorption, larger special surface area, uptake and presentation of immune cells to be triggered immune response. With these advantages NCaP had a broad prospect of application. It had been used as gene carriers for more than 30 years by way of DNA-calcium phosphate co-precipitation as an efficient method to introduce pDNA into cells. Lots of experiments showed that NCaP led to high transfection efficiency. Recently the research of NCaP as attenuated or inactivated vaccine adjuvant has achieved some results. However, until now, there are no reported studies on NCaP as peptide vaccine adjuvant. In this study, it was screened to the preparation of NCaP, and preliminary investigated NCaP’s immune effect of the CSFV peptide and Type-O FMDV peptide vaccines.1. Preparation of NCaP suspension and the quality assessment.Nano-calcium phosphate suspensions were prepared by precipitation method, and the preparation conditions were investigated and quality assessment was made. It was made with calcium chloride and dibasic sodium phosphate concentration 0.01 mol/L, Ca/P ratio 1/1 reaction 3 hours, added 50μl sodium citrate and then prepared though a high pressure homogenizer with 15000 psi for 6 cycles,0.05% CMC-Na or 0.03% HVCR as suspending agents. And well prepared nano-calcium phosphate suspensions could be made.2. Studies on the immune effect as CSFV peptide adjuvant of the NCaP.NCaP adsorpted CSFV peptide with optimal conditions in 25℃/4℃ temperature,100 μg/mL CSFV peptide concentrations and for 2 hours and the adsorption rate reached to 70%. Results of immunization experiments showed that the antibody levels of NCaP adjuvant group were lower than those of Freund’s adjuvant group and were simultaneously higher than those of free peptide. Challenge test results showed that except Freund’s adjuvant group the body temperature of rabbits in all the other groups arose, and the time of body temperature acsent of rabbits in NCaP adjuvant group postponed 16 hours compared with that in control group; Virus antigen in rabbits’ serum of Freund’s adjuvant did not be detected and those of the NCaP adjuvant group did. The studies showed that NCaP acted as CSFV peptide vaccine adjuvant could improve the immune effect to a certain extent.3. Studies on the immune effect as Type-O FMDV peptide adjuvant of the NCaP.NCaP encapsulated Type-O FMDV peptide by co-precipitation method and 1, 2-propylene glycol was used as peptide stabilizer. The mean percent rate of encapsulation efficiency reached 50%. Test showed that antibody levels of rats in NCaP adjuvant group were lower than those in oil adjuvant group and improved greatly compared with those in free FMDV peptide group. Serum cytokines test results showed that only IL-2 concentration of rats in oil adjuvant group and those in NCaP adjuvant No.2 group rose in 21 days after immunization, others had null changes. Compared with control group oil adjuvant MTT test results of NCaP adjuvant No.2 group had significant difference. NCaP acted as Type-O FMDV peptide vaccine adjuvant could improve the immune effect to a certain extent.In summary, we studied the preparation mathods of NCaP suspension and then made quality assessment. Preliminarily it was studied of the immune effects of NCaP acted as CSFV or Type-O FMDV peptide vaccine adjuvant. It was found that NCaP could improve the immune effect to some extent, but not as well as Freund’s adjuvant or oil adjuvant. |
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