A fully recombinant ELISA using in vivo biotinylated antibody fragments for the detection of potato leafroll virus

Ahmad Al-Mrabeh, Angelika Ziegler, Graham Cowan, Lesley Torrance

Research output: Contribution to journalArticlepeer-review

Abstract

A recombinant antibody fusion protein, V3HCL, which was shown previously to have specific reactivity for potato leafroll virus (PLRV), was labeled with biotin using standard chemical coupling procedures and by an in vivo method. The in vivo method proved superior giving reproducible VMCL-biotin preparations. A fully recombinant ELISA was devised incorporating V3HCL, V3HCL-biotin and streptavidin alkaline phosphatase conjugate. This assay gave comparable results for PLRV detection in potato to an assay based on immunoglobulins. The V3HCL-biotin preparations were stable and retained specific activity for more than 1 year when stored at 4 degrees C or -20 degrees C. The results demonstrate that scFv reagents derived from synthetic phage display platforms can provide effective alternatives to assays incorporating immune reagents. (C) 2009 Elsevier B.V. All rights reserved.

Original languageEnglish
Pages (from-to)200-205
Number of pages6
JournalJournal of Virological Methods
Volume159
Issue number2
DOIs
Publication statusPublished - Aug 2009

Keywords

  • ScFv
  • V3HCL
  • In vivo biotinylation
  • Recombinant antibody
  • Streptavidin
  • PLRV
  • CHAIN VARIABLE FRAGMENTS
  • PHAGE DISPLAY LIBRARIES
  • FUSION PROTEINS
  • SINGLE
  • REAGENTS
  • ENZYME
  • ASSAY

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