title: Reduced toxicity of expression, in Escherichia coli, of antipollutant antibody fragments and their use as sensitive diagnostic molecules.
abstract: Single-chain antibody fragments (scAb), specific for the chlorophenoxy acid herbicide mecoprop, have been expressed and purified from the bacterium Escherichia coli. Co-expression with the colE1-compatible, arabinose-inducible, skp expression vector pHELP1 prevented bacterial lysis and significantly increased both total and functional expression yield. The periplasmic protein, SKP, may have a role as a generic detoxification protein. Surface plasmon resonance (BIAcore 2000) analysis confirmed that the purified scAb retained similar binding kinetics to the monoclonal antibody (Mab) from which it was cloned. In competition ELISA, the bacterial scAb showed the same specificity for mecoprop and a related herbicide, MCPA, as the Mab but an increase in sensitivity for free antigen in all ELISA formats. Bacterially expressed antibody fragments provide a simple, sensitive and cost-effective alternative to the traditional production of diagnostic Mabs via tissue culture.
authors.affiliation: Department of Molecular and Cell Biology, Institute of Medical Sciences, University of Aberdeen, Foresterhill, UK.
authors.forename: G
authors.initials: G
authors.lastname: Strachan
authors.forename: S
authors.initials: S
authors.lastname: Williams
authors.forename: S P
authors.initials: SP
authors.lastname: Moyle
authors.forename: W J
authors.initials: WJ
authors.lastname: Harris
authors.forename: A J
authors.initials: AJ
authors.lastname: Porter
mesh_terms: D008456:2-Methyl-4-chlorophenoxyacetic Acid / Q000031:analogs & derivatives* / Q000032:analysis / Q000276:immunology; D000906:Antibodies / Q000235:genetics; D000918:Antibody Specificity; D001667:Binding, Competitive; D004785:Environmental Pollutants / Q000032:analysis* / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D004926:Escherichia coli / Q000235:genetics; D006540:Herbicides / Q000032:analysis* / Q000276:immunology; D007128:Immunoglobulin Fragments / Q000096:biosynthesis* / Q000235:genetics; D011485:Protein Binding; D011994:Recombinant Proteins / Q000096:biosynthesis; D012680:Sensitivity and Specificity
chemical_list: D000906:Antibodies; D004785:Environmental Pollutants; D006540:Herbicides; D007128:Immunoglobulin Fragments; D011994:Recombinant Proteins; C008340:mecoprop; D008456:2-Methyl-4-chlorophenoxyacetic Acid