Analysis of cortisol with a flow displacement immunoassay

This study investigates a flow immunoassay for cortisol based on displacement of labelled antigen analogues from immobilised antibodies. On a displacement column, two antibodies with different affinities were immobilised on protein-A Sepharose, and a construct of cortisol and horseradish peroxidase (cort-HRP) was associated to the antibodies. Perfusion of the column with cortisol displaced a proportional amount of cort-HRP. On-line detection was performed with an electrochemical detector measuring cort-HRP directly in a flow injection system. For investigating some characteristics of the displacement system, cort-HRP was also detected off-line, with a substrate for HRP, which changes OD492. We tested our system in buffer as well in serum for 2 days at a flow of 4-6 mu l min(-1). Analysis of the data show a detectable displacement signal at physiological cortisol concentrations. The specificity, stability of the columns and the influence of the antibody affinity were studied. (C) 1997 Elsevier Science S.A.