Semiquantitative, bar code version of immunochromatographic assay system for human serum albumin as model analyte

An immunochromatographic assay system was devised that can express the concentration ranges of analyte (e.g., urinary human serum albumin) as distinct numbers of the ladder bar (bar coding) for semiquantitation. We constructed a model system consisting of five membrane pad strips partially superimposed in a length. Upon wicking of sample from the bottom, the medium dissolved two different biotinylated species, antibody to the analyte and conjugates of the antibody with colloidal gold, and antigen-antibody reactions took place in the hollow space of the glass fiber membrane. After eliminating unreacted biotinylated molecules at the next strip with an immobilized albumin, the immune complexes were transferred to the pad with streptavidin immobilized in a ladder bar pattern. Analytical conditions here were set for competition between the two biotinylated species for the streptavidin binding sites. The degree of such competition was proportional to the analyte concentration and, consequently, the bar signal number was elevated as the concentration increased. Under optimal conditions for sensitivity, the analytical system responded to the analyte doses at between 30 and 120 mg/dL by producing different bar codes within 5 min.