The Experimental Study Of Different Routes In Immunization Against Dental Caries With PVAX1-SA, PVAX1-GC And PVAX1-SG In New Zealand White Rabbits

Objective:To investigate the efficiency of pVAX1-SA,pVAX1-GC and pVAXl-SG immunized in New Zealand rabbits by different routes.We further hope to explore the best effective route using in rabbit.Methods:The present study includes two parts:Test one:Identification and preparation of recombinant plasmids pVAXl-SA. pVAXl-GC and pVAX1-SG and pVAXl plasmid.First procedure in which extract a small sample of recombinant plasmids pVAX1-SA,pVAX1-GC,pVAX1-SG and vector plasmid pVAXl by Plasmid Mini Kit I. Then the recombinant plasmids were sheared by restriction endonuclease and identified by gel electrophoresis. On the basis of correct identification, pVAXl-SA,pVAX1-GC and pVAXl-SG and pVAX1 vector were isolated and purified in large quantity and were detected OD value, which were up to the requirements of the animal immunition. Test two: Immunization of New Zealand rabbits with pVAX1-SA,pVAX1-GC and pVAX1-SG. Twenty-seven New Zealand rabbits were randomly divided into three groups(A,B,C). Those groups as the model groups, and six rabbits as the controls. Every rabbit weight at 1.5kg. Each model group of rabbits were randomly divided into three groups again. A group include A1,A2,A3;B group includeB1,B2,B3;C group includeC1,C2,C3. Al group of rabbits were immunized with plasmid PVAXl-SA by intramuscular include. A2 group of rabbits were immunized with plasmid PVAxl-SA by submandibalar gland immunization. A3 group of rabbits were immunized with plasmid PVAx1-SA by nasal mucosa drip. The save method were used in other model groups, just B group of rabbits were immunized with plasmid PVAx1-GC, and C group of rabbits were immunized with plasmid PVAxl-SG. The control group of rabbits were randomly divided into two groups.In this group, half of rabbits were immunized with plasmid PVAX1 by intramuscular infection, the other rabbits were infected with by intramuscular.Each rabbit amount of plasmid DNA are 200μg(100μg/100μl).All the rabbits were immunized three times, after the first time immunity,these rabbits need interval one weeks to strengthen the immunity by the same dosage,interval two weeks to strengthen the immunity 2nd time again. Saliva samples and serum samples were collected before immunilization and after immunilization at 1,2,3,4,6,8,10weeks after immunilization, respectively. The antibody responses induced by the vaccines were detected by indirect ELISA method.Results:①On the basis of correct identification, the three vaccines were isolated and purifiedy by large QIAGEN Plasmid Giga Kit in order to meet the requirements of the study;②The level of specific antibody IgG and SIgA in serum and salivary of DNA vaccine groups were significantly elevated and continued for several weeks. However, the salivary specific antibody SIgA of intranasal immunization group and submandibalar gland immunization group was detected higher than that of the other route.Conclusions:①The recombinant plasmid pVAX1-SA,pVAXl-GC and pVAX1-SG have immunogenicity, and can provoke specific immune responses in rabbits;②The plasmid pVAX1-SG by submandibalar gland and nasal mucosa drip may be effective immunization routes.