Analytical Validation of Two Assays for Equine Ceruloplasmin Ferroxidase Activity Assessment

被引:1
|
作者
Gualandi, Stefano Cecchini [1 ]
Di Palma, Tommaso [1 ]
Boni, Raffaele [1 ]
机构
[1] Univ Basilicata, Dept Sci, Campus Macchia Romana, I-85100 Potenza, Italy
关键词
ammonium iron(II) sulfate; blood plasma; ceruloplasmin; enzyme; horse; o-dianisidine; oxidase activity; ACUTE-PHASE PROTEINS; SERUM CERULOPLASMIN; OXIDATIVE STRESS; OXIDASE ACTIVITY; LIPID-PEROXIDATION; AUTOMATED-ASSAY; BIOMARKERS; PLASMA; COPPER; COWS;
D O I
10.3390/vetsci10100623
中图分类号
S85 [动物医学(兽医学)];
学科分类号
0906 ;
摘要
Ceruloplasmin (Cp) assessment in biological samples exploits the oxidase activity of this enzyme against several substrates, such as p-phenylenediamine (p-P), o-dianisidine (o-D) and, most recently, ammonium iron(II) sulfate (AIS). Once developed in humans, these assays are often used in veterinary medicine without appropriately optimizing in the animal species of interest. In this study, two assays using AIS and o-D as substrates have been compared and validated for Cp oxidase activity assessment in horse's plasma. The optimization of the assays was performed mainly by varying the buffer pH as well as the buffer and the substrate molar concentration. Under the best analytical conditions obtained, the horse blood serum samples were treated with sodium azide, a potent Cp inhibitor. In the o-D assay, 500 mu M sodium azide treatment completely inhibits the enzymatic activity of Cp, whereas, using the AIS assay, a residual analytical signal was still present even at the highest (2000 mu M) sodium azide concentration. Even though the analytical values obtained from these methods are well correlated, the enzymatic activity values significantly differ when expressed in Units L-1. A disagreement between these assays has also been detected with the Bland-Altman plot, showing a progressive discrepancy between methods with increasing analytical values.
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页数:11
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