Preparation Of McAbs Against Immunodominant Fragment Of S.aureus IsdB And Primary Analyses Of Eptitopes To The McAbs

Staphyloccocus aureus infection has become a big problem worldwide, and the harm to human is more serious due to the appearance and spread of resistant strains. Vaccine against S.aureus is urgently needed. Wide studies showed traditional vaccines, such as attenuated live vaccine and inactivated vaccine, capsule polysaccharides vaccine, and toxoid vaccine were not available for S.aureus, some surface proteins of S.aureus were ideal potential vaccine. Iron-regulated surface determinant B (IsdB), demonstrated a good immunogenicity, but IsdB epitopes inducing immuno-protection are not yet clear.Recombinant IsdB3 (129-361aa) was purified and used as an immunogen for BALB/c mice in this experiment. Six cell strains, which could stably secret monoclonal antibodies (McAbs) against IsdB3, were obtained by hybridoma technique, and named 2E1,2E3,3A7,3B6,3H5 and 4D3. The chromosome number of the cell strains were between 80-96 detected by Giemsa's stain method, consistent with fusion cell genetic algorithm. The heavy chain subtype of 2E1,2E3 and 3H5 McAbs were IgG1,3B6 and 4D3 were IgG2b, and 3A7 was IgG2a. All light chains of the McAbs wereκ. There was no cross reaction between IsdB3 and TraP detected by the McAbs. This indicated the McAbs were of good specificity. All 6 McAbs could specifically bind to IsdB3 by Western blot, but 3A7,3B6 and 4D3 could bind to S.aureus by indirect ELISA based on coating with whole S.aureus.Five pairs of primer were designed according to the sequence of IsdB3. Chromosome DNA of S.aureus Newman strain was used as the templet to amplify isdB3a, isdB3b, isdB3c, isdB3d and isdB3e fragments. The expressed peptides coded by these fragments were analyzed with McAbs 3A7,3B6 and 4D3 respectively by Western blot. All expressed peptides could bind to the 3 McAbs. This result indicated that the IsdB3 antigen epitopes exposed on bacterium surface were mimic epitopes. Using phage display technique, the phages interacting with 3B6 McAbs were screened in heptapeptide library and the peptide sequence of the phages were analyzed. The result further confirmed that the IsdB3 antigen epitope exposed on bacterium surface was a mimic epitope. This research would provide important reference for further research on anti-S. aureus poly antigen chimeric epitope vaccine.