Usefulness of ferroxidase activity of ceruloplasmin in the diagnosis of Wilson’s disease

Abstract

Serum ceruloplasmin is one of the most commonly used screening tests for Wilson’s disease. However immunological assays for ceruloplasmin are not recommended for diagnosis and management of Wilson’s disease through calculation of free copper index. Enzymatic methods using non-physiological substrates have toxicity and stability problems, making them difficult to automate. Ferroxidase assays may be a satisfactory alternative for measuring serum ceruloplasmin. The o-dianisidine hydrochloride manual method for estimation of serum ceruloplasmin enzyme activity was compared with an automated method using the ferroxidase activity of ceruloplasmin in measurement in a double blind study in 91 consecutive patients screened for Wilson’s disease. The o-dianisidine and ferroxidase methods both successfully identified 7 patients with Wilson’s disease. Values for these 7 patients in the o-dianisidine and ferroxidase methods were median 5.0 (range 0–16.0 U/L) and median 45.0 (range 4–166 U/L) respectively. There were 7 other positive values (<62 U/L) with the o-diansidine method and 2 (<200 U/L) with the ferroxidase method, where WD was not confirmed. ROC curves for both methods showed area under the curve of 0.998 for o-dianisidine and 0.997 for ferroxidase. Using literature cut off values of 62 U/L and 200 U/L respectively both methods had 100% sensitivity and specificity was 91.7% (o-dianisidine) and 97.6% (ferroxidase). For the o-dianisidine assay, specificity was improved to 98.8% using a cut off of 22.5 U/L. In the 84 persons (46 adults and 38 children) in whom the diagnosis of Wilson’s disease was not established, the mean value for ceruloplasmin activity by the o-dianisidine and ferroxidase methods was 124.7 ± 48.7 U/L and 571.4 ± 168.1 U/L respectively. There were no significant differences between sex or age of patients (p > 0.29). In a subsequent evaluation with 372 specimens, the Pearson correlation coefficient between the assays was 0.908, p < 0.01, slope 4.06, intercept 265.8, with the manual assay as the x-axis. The ferroxidase assay is a suitable replacement for the o-dianisidine assay in detecting patients with Wilson’s disease.