Dual-column immunoassays using protein G affinity chromatography.
Tandem protein G affinity and reversed-phase chromatography (RPC) columns, coupled with a switching valve, were used for on-line immunoassays of antibodies and antigens. Columns with reversibly immobilized antibodies were prepared by adsorbing antibodies on the protein G column. Following antigen capture in the immunoaffinity column, antigen-antibody complexes were desorbed, dissociated, and transferred to the RPC column where they were separated and quantified. This system was used to determine the titer of a rabbit anti-human transferrin antibody sample with a precision of +/- 2%. Quantitation of human transferrin in human serum had a precision of +/- 6% and showed good agreement with rate nephelometry. The linear dynamic range for the transferrin, antigen immunoassay was 5 x 10(1) to 1 x 10(5) ng with a precision of +/- 3.5%.
Janis
L J
LJ
Regnier
F E
FE
D002846:Chromatography, Affinity; D007118:Immunoassay / Q000295:instrumentation*; D009419:Nerve Tissue Proteins / Q000032:analysis*
C029152:G-substrate; D009419:Nerve Tissue Proteins