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Development of a Rapid Capture ELISA
Using
PCR Products and the PinPointTM System
By Ingrid G. Winkler, Ph.D., School of Pharmacy and Medical
Science, University of South Australia, Adelaide, Australia
Jean-Pierre Levesque, Ph.D., Haematology Division, Hanson Centre for Cancer Research,
Adelaide, Australia
Robert L.P. Flower, Ph.D., Pacific Laboratory Medicine Services, Royal North Shore
Hospital, St. Leonards, Australia
A simple procedure for the development of an ELISA is presented. The
procedure uses the PinPointTM Xa-1 T-Vector System to express a protein antigen encoded by a
PCR product. Because the PinPointTM System adds a biotin tag to the
protein, a single-step purification by affinity for streptavidin allows direct use in
ELISA. We report the use of this procedure to express a recombinant protein from the
nucleocapsid domain of the feline foamy virus (FeFV) gag gene, fused with a biotin tag.
This fusion protein was applied directly to streptavidin-coated ELISA wells. Antibody to
FeFV was detected in this ELISA with a 100% correlation to other detection methods,
including immunoblot, serum neutralization and virus isolation.
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