A labeling method and purely colorimetric immunoassay based on poly-DL-lysine and a pigmented copper cluster

A non-biological binding action between a biotinylated, alpha-linked poly-DL-lysine (PDLL) and a pigmented, red-violet cluster anion of copper and penicillamine (RVC) may be employed as an affinity labeling method. The neutral pH association constant of this interaction is on the order of 10⁵ M^-1, and the complex survives zone electrophoresis for more than 40 min in an acetate buffer (pH 5), migrating as a cation and dissociating very slowly. Using spectrophotometric measurements at 520 nm, the RVC/PDLL binding interaction is the basis of a purely colorimetric immunoaffinity assay with a sensitivity threshold of approximately 0.3 nmol (as antibody or antigen). The method, which is based on differential elution rates, could be scaled down for capillary methods with increased sensitivity. This labeling application uses PDLL in the 30 000-70 000 g/mol molecular weight range. (C) 2000 Elsevier Science B.V.