Preparation and characterization of digoxin antibody and its application to immunoassays:comparison of performance characteristics between enzyme immunoassay and immunostrip test

摘要

After characterizing digoxin immunogens and antibodies,we investigated the assay performances of enzyme immunoassay(EIA),biotin-streptavidin mediated enzyme immunoassay (B-Av EIA),and immunostrip test and compared the effectiveness of these tests for detecting the presence of digoxin and its analogues.The development of a rapid and simple detection device has been needed.In our study,we deterined digoxin levels by EIA and B-Av EIA,and compared these tests ith the immunotest strip to determine the possiblity of personal monitoring at the bedside.Three kinds of digoxin antibodies were produced from various digoxin-BSA immunogens(digoxin/BSA molar ratio of 15:1 ,50:1, and 200:1 in their preparation step).Teh antibodies produced were purified by the immunoaffinity chromatography using digoxin-KLH or digoxin-BSA as the affinity ligand.Antibody #7 and its matching coating ligand of digoxin-KLH were selected by titration and sensitivity studies for use in EIA and B-Av EIA.Colloidal gold-labeled antibody #7 on a glass fiber membrane and digoxin-BSA on a nitrocellulose membrane were selected for the immunostrip preparation.The EIA adn B-Av EIA culd detect 0.01 ppb digoxin within 2.5 h,but the immunostrip could detect 0.01 ppm vithin 3 min,which was three orders less sensitive than EIA or B-Av EIA.