Preparation and preliminary application of monoclonal antibody against biofilm-associated protein of Acinetobacter baumannii
Objective To prepare monoclonal antibody(mAb)against biofilm-associated protein(Bap)of Acinetobacter baumannii and to initially develop an immunological detection technology for A.baumannii.Methods The Bap gene sequence was synthesized chemically,expressed by prokaryotic expression system and then purified by affinity chromatography to obtain recombinant protein Bap-His.The spleen cells of two female BLAB/c mice immunized with Bap-His recombi-nant protein were fused with SP2/0 myeloma cells and screened for the positive hybridoma cell lines by indirect ELISA.The monoclonal antibodies were prepared,analyzed by SDS-PAGE and identified by Western blot.Based on indirect ELISA,the immunological detection technology for A.baumannii was developed by using the obtained monoclonal antibodies,and the specificity and lowest detection limit were verified.Results Two hybridoma cell lines were screened,and two monoclonal antibodies,mAb-2F and mAb-7E,were prepared.Both monoclonal antibodies had the purity of over 90％,and showed specific binding with the natural protein of A.baumannii.The immunological detection technology developed by two monoclonal anti-bodies only cross-reacted with A.baumannii,but did not with other respiratory pathogens.The lowest detection limits were 1.6 × 105 and 8.0 × 104 cfu/mL respectively.Conclusion The prepared Bap monoclonal antibody has high purity and can provide biological materials for the study of Bap related functions;the initially developed immunological detection technology for A.baumannii has good specificity and sensitivity,which provides technical support for the detection of A.baumannii.
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