NON-INSTRUMENTED ENZYME IMMUNOASSAY: GENERAL METHOD UTILIZING KINETIC BINDING EFFECTS

摘要

A method for detecting the presence or amount of an analyte in a sample by means of an enzyme-labelled immunoassay that utilizes an enzyme-labelled competitive binding compound that competes with the analyte for binding to an anti-analyte antibody specific for the analyte, in which the improvement comprises utilizing as the competitive binding compound a derivative of the analyte comprising an analyte moiety, a bulky immunogenic moiety bound to the analyte moiety at a location on the analyte used to attach an immunogen to the analyte when preparing the anti-analyte antibody, and an enzyme reporting group bound to the bulky linking moiety. The bulky linking group and the enzyme are selected so that together they are of sufficient size to slow diffusion of the competitive binding compound and reduce its ability to bind with the anti-analyte antibody (relative to analyte affinity) by kinetic effects in an amount sufficient to steepen the competitive binding curve for the assay, so that a sharper positive/negative threshold can be distinguished in a non-instrumented enzyme immunoassay without requiring structural manipulations at the antibody binding site.