|
Abstract
|
Background: Pseudomonas aeruginosa is the leading cause of nosocomial infections, especially in people with a
compromised immune system. Targeting virulence factors by neutralizing antibodies is a novel paradigm for the
treatment of antibiotic-resistant pseudomonas infections. In this respect, exotoxin A is one of the most potent
virulence factors in P. aeruginosa. The present study was carried out to identify a novel human scFv antibody
against the P. aeruginosa exotoxin A domain I (ExoA-DI) from a human scFv phage library.
Methods: The recombinant ExoA-DI of P. aeruginosa was expressed in E. coli, purified by Ni-NTA column, and used
for screening of human antibody phage library. A novel screening procedure was conducted to prevent the
elimination of rare specific clones. The phage clone with high reactivity was evaluated by ELISA and western blot.
Results: Based on the results of polyclonal phage ELISA, the fifth round of biopanning leads to the isolation of
several ExoA-DI reactive clones. One positive clone with high affinity was selected by monoclonal phage ELISA and
used for antibody expression. The purified scFv showed high reactivity with the recombinant domain I and fulllength native exotoxin A.
Conclusions: The purified anti-exotoxin A scFv displayed high specificity against exotoxin A. The human scFv
identified in this study could be the groundwork for developing a novel therapeutic agent to control P. aeruginosa
infections
|