A combined vaccine of Equine influenza (EI), equine herpesvirus-1 (EHV-1), rabies virus and tetanus toxoid adjuvanted with saponin and Alhydrogel was prepared. Quality control testing of such vaccine revealed that it was free from foreign contaminants, safe for Guinea-pigs, mice and mares. It was inoculated into Guinea -pigs as a preliminary evaluation for its potency and it was proven safe and potent. Also, it exceeded the permissible protective level allowed to be used for the vaccination of horses. Two groups of mares were used for evaluating the potency of the vaccine. The first one received two doses (5 ml) of the prepared vaccine with 4 weeks interval while the 2 nd group was kept without vaccination as control. The mean haemagglutination inhibition (HI) influenza antibody titer reached its maximum at the 2 nd month post-vaccination (MPV); 2048 while EHV-1 antibodies reached the peak at the 3 rd MPV as recorded by ELISA and neutralizing indices; 1790 and 3.50. Rabies antibodies were detectable in vaccinated mares by the 2 nd week post-vaccination showing titers of 16 and 1.0 by serum neutralization test (SNT) and ELISA, respectively recording their peaks (128 by SNT and 2.15 by ELISA) at the 2 nd MPV. Tetanus antitoxic titer increased till reaching the peak at the 2 nd MPV (40IU/ml) as determined by toxin neutralization test. Depending on these results, it could be concluded that the prepared inactivated EI, EHV-1, rabies and tetanus vaccine is safe and potent for mares providing them with good protective levels of specific antibodies against the 4 used antigens up to 6 months.

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In this study, neutralizing antibody response in Guinea pigs and mice inoculated by African horse sickness (AHS) bivalent inactivated oil adjuvant vaccine was similar when compared serologically with that in horse. Therefore, using G. pigs and mice may be a method of great value in potency test for routine immunization. Potency test of the prepared vaccine using an immunization challenge protocol in adult mice was studied, using challenge virus with 100, 50 and 25 LD 50 . The highest protection level (95%) was with the dilution of 50 LD 50 /0.05 ml.

Vaccination remains an important strategy to fight Equine influenza (EI), Equine herpesvirus-1 (EHV-1) and tetanus infection, in combination with management measures. A combined inactivated EIV, EHV-1 and Tetanus toxoid (TT) vaccine adjuvanted with saponin and Alhydrogel was prepared and tested in four groups of guinea pigs in comparison with each vaccine separately beside another group as control.The humoral immune response in vaccinated guinea pigs against EI, EHV-1 and tetanus was evaluated by Haemagglutination Inhibition (HI) antibody titers, Enzyme linked immunosorbent assay (ELISA) titers, and toxin neutralizing (TN) antibody titers respectively. All vaccines proved to be safe and potent in guinea pigs. Also, the obtained antibodies titers with combined vaccine was nearly similar to titer obtained with separated vaccine of each target agent in vaccinated guinea pigs. The evaluation of keeping quality of the prepared combined inactivated vaccine showed that it is was stable at 4C up to 12 months and for 3 months at room temperature.

The use of vaccines in the prevention and control of infectious diseases in the horse is a widely accepted practice. A combined inactivated EIV, EHV-1 and tetanus toxoid vaccine adjuvanted with alhydrogel and saponin was evaluated in horses. Two groups of horses, group A was inoculated with two doses of the combined vaccine one month apart and revaccinated at 6 months and group B was kept as control. The immune response of vaccinated horses against EI, EHV-1 and tetanus was evaluated by measuring Haemagglutination Inhibition (HI), Enzyme linked immunosorbent assay (ELISA), Complement Fixation (CF) and toxin neutralizing antibody titers (TN). Vaccinated horses had high antibody titers, above the level for clinical protection against influenza and tetanus until 12 months and 6 months for EHV-1 after the third vaccination. A vaccination scheme consisting of three doses with combined inactivated EI, EHV-1 and TT vaccine adjuvanted with saponin and alhydrogel, two doses at one-month interval and Revaccination after 6 months, Followed by alternating vaccinations with monovalent inactivated EHV-1vaccine and combined inactivated EI-EHV& TT vaccine every 6 months is proposed to maintain maximal protection against equine influenza and herpesvirus and tetanus infections.