Production of ScFv antibody fragments following immunization with a phage-displayed fusion protein and analysis of reactivity to surface-exposed epitopes of the protein F of Pseudomonas aeruginosa by cytofluorometry. Academic Article uri icon

Overview

abstract

  • To increase the possibilities of obtaining antibodies to surface-exposed epitopes of Pseudomonas aeruginosa protein F, we immunized mice with cloned and expressed oprF gene as a gIII-fusion protein displayed on the M13 phage surface. The fusion protein elicited mouse antibodies reacting with the purified protein F at a limit dilution of 1:10,000. Recombinant clones expressing antibody fragments were constructed from the genes of selected B cells of hyperimmunized mouse after a first round of panning against the protein F. Expression of single chain Fv (ScFv) antibody fragments to the protein of P. aeruginosa was detected by ELISA in 20 of 384 clones obtained after the first panning selection. The 20 positive clones recognizing different protein F epitopes as demonstrated by ELISA were assayed by flow cytometry to identify antibody fragments reacting only with surface-exposed epitopes of the protein F on whole bacteria; one of the 20 clones tested showed a level of reactivity compatible with surface-exposed epitope that can lead to ulterior developments in targeting studies.

publication date

  • August 1, 1995

Research

keywords

  • Epitopes
  • Immunoglobulin Fragments
  • Immunoglobulin Variable Region
  • Porins
  • Pseudomonas aeruginosa
  • Recombinant Fusion Proteins

Identity

Scopus Document Identifier

  • 0029122181

Digital Object Identifier (DOI)

  • 10.1089/hyb.1995.14.323

PubMed ID

  • 8522342

Additional Document Info

volume

  • 14

issue

  • 4