New enzymatic assay for calcium in serum

Area of Science:

• Biochemistry
• Clinical Chemistry
• Enzymology

Background:

• Accurate serum calcium measurement is crucial for diagnosing and managing various medical conditions.
• Existing methods may have limitations in terms of speed, complexity, or susceptibility to interference.
• Development of novel, efficient, and reliable calcium assays is an ongoing area of clinical chemistry research.

Purpose of the Study:

• To establish a simple, rapid, and reliable kinetic assay for quantifying calcium in serum.
• To utilize urea amidolyase (EC 3.5.1.45) from yeast species as the core enzyme for calcium detection.
• To validate the performance characteristics and assess the interference profile of the newly developed assay.

Main Methods:

• A kinetic assay was developed based on the inhibitory effect of calcium on yeast urea amidolyase.
• Endogenous ammonium ions were removed using glutamate dehydrogenase (GLDH; EC 1.4.1.4).
• Ammonium ion production, inversely proportional to calcium concentration, was measured spectrophotometrically at 340 nm after NADP+ formation catalyzed by GLDH.

Main Results:

• The assay demonstrated good precision with within-run coefficients of variation (CVs) of 1.7-3.2% and day-to-day CVs of 2.8-4.1%.
• Analytical recovery ranged from 92-112%, indicating accuracy.
• The assay showed excellent correlation with established methods (o-cresolphthalein complexone and another enzymatic method), with correlation coefficients (r) of 0.949 and 0.955, respectively.

Conclusions:

• The developed kinetic assay using urea amidolyase provides a simple, rapid, and accurate method for serum calcium determination.
• The assay is robust, with minimal interference from common serum components and substances.
• This novel enzymatic method offers a reliable alternative for routine clinical laboratory analysis of calcium.