caa a22 4500 397494203 CHVBK 20180308164516.0 cr unu---uuuuu 161202e199509 xx s 000 0 eng 10.1093/oxfordjournals.jbchem.a124933 doi (NATIONALLICENCE)oxford-10.1093/oxfordjournals.jbchem.a124933 Production of a Paraquat-Specific Murine Single Chain Fv Fragment [Elektronische Daten] [Cecilia M. Devlin, Mark R. Bowles, Ross B. Gordon, Susan M. Pond] Producing an effective antidote against poisoning by the herbicide paraquat (PQ) has proven to be an elusive goal. One approach that holds some promise is immunotherapy with antibody fragments. In this study we detail the production of a single chain Fv fragment (scFv) specific for paraquat by linking cloned heavy (VH) and light chain (VI) variable region genes via the peptide spacer (Gly4-Ser)3. These genes were obtained from hybridoma cells secreting a PQ-specific murine monoclonal antibody. The scFv (28 kDa) was expressed at 4% of the total cell protein by the Escherichia coli vector, pPOW, but was associated with the membranes. After solubilisation and reduction, the scFv was renatured by rapid dilution. Western blotting confirmed that the refolded scFv had similar structural properties to the parental mAb. The isoelectric point of the scFv (7.0) is equal to the value calculated from the deduced amino acid sequence. Surface plasmon resonance was used to demonstrate specific PQ binding by the refolded scFv (Ka=1.24x106 M-1) which is similar to that determined for the parent Fab fragment (4.6X 106 M-1). © by the Journal of Biochemistry Regular Papers nationallicence murine antibody nationallicence paraquat (1, 1‘-dimethyl-4, 4'-bipyridinium dichloride) nationallicence protein refolding nationallicence single chain antibody nationallicence surface plasmon resonance nationallicence Devlin Cecilia M. University of Queensland, Department of Medicine, Princess Alexandra Hospital, Brisbane, QLD, 4102, Australia aut Bowles Mark R. University of Queensland, Department of Medicine, Princess Alexandra Hospital, Brisbane, QLD, 4102, Australia aut Gordon Ross B. University of Queensland, Department of Medicine, Princess Alexandra Hospital, Brisbane, QLD, 4102, Australia aut Pond Susan M. University of Queensland, Department of Medicine, Princess Alexandra Hospital, Brisbane, QLD, 4102, Australia aut The Journal of Biochemistry Oxford University Press 118/3(1995-09), 480-487 0021-924X 118:3<480 1995 118 jbchem https://doi.org/10.1093/oxfordjournals.jbchem.a124933 text/html Onlinezugriff via DOI 1 other jats NATIONALLICENCE 856 40 https://doi.org/10.1093/oxfordjournals.jbchem.a124933 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Devlin Cecilia M. University of Queensland, Department of Medicine, Princess Alexandra Hospital, Brisbane, QLD, 4102, Australia aut NATIONALLICENCE 700 1- Bowles Mark R. University of Queensland, Department of Medicine, Princess Alexandra Hospital, Brisbane, QLD, 4102, Australia aut NATIONALLICENCE 700 1- Gordon Ross B. University of Queensland, Department of Medicine, Princess Alexandra Hospital, Brisbane, QLD, 4102, Australia aut NATIONALLICENCE 700 1- Pond Susan M. University of Queensland, Department of Medicine, Princess Alexandra Hospital, Brisbane, QLD, 4102, Australia aut NATIONALLICENCE 773 0- The Journal of Biochemistry Oxford University Press 118/3(1995-09), 480-487 0021-924X 118:3<480 1995 118 jbchem Metadata rights reserved CC BY-NC-4.0 http://creativecommons.org/licenses/by-nc/4.0 nationallicence BK010053 XK010053 XK010000 NATIONALLICENCE NATIONALLICENCE NL-oxford