Preperation And Immunologic Evaluation Of Inactived Rotavirus And Inactived Hepatitis A Combined Vaccines

As early as at the end of 1980s,WHO suggested that Using combined vaccine is a effective way to Prevent diseases when they formulated childhood vaccines immunization(CVI) programs. Therefore, the development of combined vaccine is not only currently trend, but also the direction of future vaccine research. Combined vaccines not only solve the economic burden,but also reduced the mental burden. It is efficient to solve these problems by reducing single product numbers and vaccination numbers. At present there are some combined vaccines for our immune program, such as pertussis diphtheria tetanus mixed vaccine, measles parasynanche rubella mixed vaccine, Hepatitis A hepatitis B mixed vaccine, etc. Now in our country there no combined vaccine based with rotavirus.Human rotavirus (human rotavirus, HRV) is the most important pathogen of severe diarrhea in infants and children under 5 years old worldwide, HRV infection is the main cause of death from infantile diarrhea. Each year about 60 million infants and children died of an RV infections diarrhea. At present the use of live attenuated RV vaccine in preventing an RV infection plays a certain role, but the vaccine may caused side effects such as extrinsic factor pollution and the reversion of RV virulence. Up to now,the problem has not been solved perfectly.World Health Organization (WHO) has been actively promoting research and development of different forms of rotavirus vaccines. Inactivated RV vaccinea is encouraged to develop to to prevent and control HRV infection. At present we have finished laboratory scale inactivated vaccine preparation,it is the basis of developing combined vaccine.Viral hepatitis is caused by a variety of hepatitis virus mainly in acute liver damage or persistent infection systemic and has been found all over the world. In hepatitis patients 30～50% belong to armour hepatitis. It is associated with social and economic development level, closely related to environmental hygiene standards. Developed countries infection rates of up to 50-1,000/10 million, developing country for 100 to 300/10 million. In most regions of China belongs to hepatitis a high popular area. Due to the use of hepatitis A combined vaccines,the quantity has been greatly reduced in the popular and incidence of hepatitis A. inactived hepatitis A vaccine (IHAV) play an important role.If we can use two or more effective vaccine with different proportion vaccine to obtain combined vaccine,it will lighten the by the economic and spirit of burden. Based on this requirement,we considering that the IRV and IHAV could be mixed to obtain combined vaccine.Because IRV and IHAV be of the same physical and chemical properties, be vaccinated to the same immune object and each vaccine immunization procedure should basically be same, we mixed the inactived rotavirus vaccine (IRV) P[2]G3 strain and inactived hepatitis A vaccine (IHAV) to obtain combined vaccine.Meanwhile, this study also carried out the immunogenicty and protective effect of IRV P[2]G3 strain and IHAV vaccine (IHAV) combined vaccines in mice.The first step of this study:the preparation of combined vaccines.Experimental methods:P[2]G3 strain rotavirus prepared in a large scale were purified by centrifugation, microfiltration, ultrafiltration and chromatography. Purified viruses were infectious and showed typical feature of the rotavirus with structural integrity. The genomes of purified rotavirus had no change. Purified rotavirus were completely inactivated when final concentration 0.00925% formaldehyde was added into virus fluids, following heat for 30min at 56, and then continued to incubate for 72 hours at 37℃.It has been proved that thre is no residual virulence by three generations blind passage,the virus is completely inactivated. Accurate antigen definitation was made to decide how much dose of vaccines should be given for mice,then we can use the inactived P[2]G3 rotavirus was adsorbed with aluminum hydroxide adjuvant mixed IHAV vaccine to have the combined vaccines.The second step of this study:The immunogenicty and protective effect of combined vaccines.Methods:the mice were divided into blank group,control group and test group.the combined vaccine is vaccinated intramuscularly three times to mice by the hind legs.the immunization is be separated by two weeks,and then to detect the levels of serum and fecal rotavirus specific antibody and CD4+T cells and CD8+Tcells after immunization with IRV and IHAV in mice.Results:By subcutaneous injection,after two weeks, the antibody titers of SA11 stain serum IgG could be tested.after four weeks, the antibody of hepatitis A could be tested. after six weeks, antibody level to reach peak.the antibody titers of SA11 stain serum IgG achieve 160412.33±24123.54. the antibody titers of SA11 stain intestinal IgG achieve4.76±0.72. the antibody titers of SA11 stain neutralizing antibodies achieve10240. the antibody titers of hepatitisA achieve 28.50876±6.53. The percentage of the CD4+T cells achieve 14.52±3.83 and CD8+T cells achieve 3.9±0.72.By t test,P<0.05.it was found that the combined vaccines induced not only humoral immune responses but also CD4+T lymphocyte responses.Some science foundation were providee for IRV and 1HAV combined vaccines by This reaserch.