Immunoprophylactic potential of anti-idiotype foot-and-mouth disease vaccine

Abstract

The present study was designed to develop an anti-idiotype foot-and-mouth disease antigen for the evaluation of immunoprophylactic potential. Three serotypes of foot-and-mouth disease virus were procured from the veterinary research institute, Peshawar. Re-characterization of the virus was done through PCR and further serotyping was performed by indirect sandwich ELISA (IsELISA). The inactivated three serotypes of the virus were injected in goats for the preparation of idiotype antibodies (IgG). The goat serum containing 90% idiotype antibody titre was processed for separation of idiotype antibodies through Octanoic acid-ammonium sulfate precipitation method. Idiotype was purified and the fragment antibody binding (Fab) component was separated through pepsin digestion that was analyzed through SDS-PAGE. The protein concentration of Fab was adjusted to 5 mg/mL and 10 mg/mL. Fab of IgG was emulsified in MONTANIDE™ ISA 206 VG and the prepared idiotype antigen suspension was injected into two groups of the layer birds. The eggs were collected and the presence of anti-idiotype antibodies in the egg yolk was confirmed through agar gel immunoprecipitation test (AGPT). The diluted egg yolks were processed for the separation and purification of anti-idiotype antibodies through ammonium sulfate precipitation technique. The purified IgY proceeded to pepsin digestion and Fab component was obtained. Fab of anti-idiotype antibody, after digestion, were analyzed through SDS-PAGE and the protein concentrations were adjusted to 10 mg/mL. The Fab of IgY was emulsified in Montanide (1:1) and the anti-idiotype FMD antigen was injected as a surrogate antigen for FMD virus in mice and calves. Sterility, safety and stability studies of anti-idiotype FMD antigen were performed. Immune response of Montanide adjuvanted monovalent and trivalent anti-idiotype FMD antigen was determined in mice. The comparative immune response of Montanide adjuvanted trivalent anti-idiotype FMD antigen and trivalent FMD commercial vaccine was done in mice and calves. The comparative mean antibody titre was determined through least significant difference followed by factorial analysis. Foot-and-mouth disease virus was detected through PCR at 131 bp. Is-ELISA confirmed three serotypes of Foot-and-Mouth Disease virus, Asia 1, A and O, at OD value < 0.1 at 1/10th dilution. The AGPT results indicated early development of anti-idiotype antibodies in layer birds injected with 10 mg/mL Montanide adjuvanted idiotype antigen. The development of antiidiotype antibodies in egg yolk was detected within 18 hrs of incubation on day 14 from the last injection. At day 45 post-immunization (PI) in mice, Montanide adjuvanted monovalent antiidiotype FMD antigens produced initial antibody titre of 78.80%, 81.30% and 81. 20% for serotype A, Asia 1, and O respectively. The antibody titre decreased to 78.10%, 79.50% and 78.90% respectively at day 60 PI. The more stable immune response against serotype A was recorded at day 60 PI. Montanide adjuvanted trivalent anti-idiotype FMD antigen in mice produced highest antibody titre of 81.60% at day 45 compared to Montanide adjuvanted FMD vaccine that produced titre of 77.50% at day 45 PI. A slow decrease of 1-2% in antibody titre of Montanide adjuvanted trivalent anti-idiotype FMD antigen in mice at day 60 was recorded. The immune response of Montanide adjuvanted trivalent anti-idiotype FMD antigen in calves was persistently 80% while titre of Montanide adjuvanted FMD vaccine decreased to 74% at day 60 PI.