Induction of bovine respiratory syncytial virus vaccinal immunity
[Thesis]
M. Zulfiqar
M. L. Kaeberle
Iowa State University
1995
158
Ph.D.
Iowa State University
1995
Electron microscopic examination of ultrathin sections of BRSV strain 375-infected Vero cells revealed not only spherical and filamentous forms of the virus but also bridges between the spherical particles. The bridges did not appear to be arranged at a particular angle. Because of the bridges the virus particles occurred in large aggregates requiring that this factor be taken into account while purifying the virus. Most of the observed mature particles were extracellular. However a few spherical particles with and without internal structure were also present in intracytoplasmic vesicles. A panel of monoclonal antibodies (MAbs), derived from the fusing of the primed BALB/C mice splenocytes to SP2/O cells, were generated against the N protein of BRSV. The MAbs were characterized by ELISA, a radioimmunoprecipitation assay, indirect immunofluorescence, and epitope mapping. According to the class and subclass of Ig these MAbs can be grouped into four groups. Eight MAbs, two from each group, were used for epitope mapping. The reaction pattern of the MAbs indicated 4 to 6 antigenic sites on the N protein. Two monoclonal antibodies 65-12 (IgM) and 38-6 (IgG2a) were used to make solid matrix-antibody-antigen complexes (SMAA). The SMAA complexes so formed were washed five times with PBS, suspended in an equal quantity of aluminum hydroxide gel and used for immunization of two groups of lambs. In immunized groups the maximum immune response in terms of lymphocyte blastogenic responses was observed on day 11 postvaccination. However, the response of animals in group B, vaccinated with SMAA complexes containing MAb (IgM) was significantly higher than the group C, vaccinated with SMAA complexes containing MAb (IgG2a). The group B animals also exhibited a marked humoral response to N protein in western immunoblot assay as compared to group C and control animals 11 days postvaccination. None of the animals developed a SN antibody titer after vaccination. Subsequent challenging, 21 days postvaccination, with heat inactivated BRSV resulted in a far better SN titer in the SMAA complexes vaccinated groups than the control group. Not only did vaccinated animals developed a higher titer but the response developed earlier since all vaccinated animals seroconverted within five days of challenge.