Automated bichromatic analysis of serum ceruloplasmin

An automated micromethod is described for measurement of serum ceruloplasmin by assay of its p-phenylenediamine oxidase activity using the Abbott bichoromatic analyzer. Ten mul of serum are mixed with 250 mul of p-phenylenediamine (PPD) substrate (9.2 mmole per liter) in acetate buffer (0.1 mole per liter, pH 5.45). Spectrophotometric measurements of the rate of formation of the purple oxidation porduct of PPD are performed after a 10 min delay for thermal equilibration at 37 degrees and for avoidance of the lag-phase of the enzymatic reaction. The coefficients of variation of replicate analyses of normal serum by this technique are 1.1 percent (within-the-run) and 3.3 percent (day-to-day). Measurements of ceruloplasmin concentrations in serums from 75 patients by this automated method provided close correlation with measurements by a manual reference procedure (correlation coefficient=0.973). The mean concentration of ceruloplasmin in serums from 64 healthy men was 29 mg per dl (central 95th percentile limits=22 to 40 mg per dl).