Production, Characterization and Purification of Monoclonal Antibody Against Acinetobacter Baumannii

Background: Pleomorphic Acinetobacter baumannii is a highly successful nosocomial pathogen. It is an obligate aerobic, gram negative bacteria that causes nosocomial pneumonia and meningitis among hospitalized patients. Broad range resistance to multiple antibiotics has been the biggest challenge to treat A. baumannii infection. Here, we described the production, characterization, purification and identification of monoclonal antibody specific against the multi-drug resistant A. baumannii. Methods & Materials: BALB/c mices were immunized with formaldehyde-fixed A. baumannii M28-47 strain. The spleen was removed and fused with myeloma cells. A series of monoclonal antibodies (mAb) specific to A. baumannii were produced. Characterization of the mAb was performed using enzyme-linked immunosorbent assay (ELISA) with subclass specific goat antisera. Antibodies were purified by using the protein G-linked magnetic beads. SDS-PAGE and immunoblot were performed to confirm the presence of the heavy and light chain of IgG of the purified antibody. Mass spectrometry was used to further confirm the identity of the antibody. Immunoblotting was performed to detect the A. baumannii protein. Results: All the hybridoma clones produced IgG1 subclass antibodies. SDS-PAGE of the purified hybridoma antibody showed the presence of the antibody heavy chain and light chain. The presence was also confirmed by ELISA, immunoblot and mass spectrometry. All the purified antibodies detected A. baumannii protein prepared on nitrocellulose membrane from A. baumannii cell lysate. Conclusion: Monoclonal antibodies specific against A. baumannii was successfully prepared. This mAb could be useful as reagents for studying A. baumannii.