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Updated: Jun 23, 2026
A Semi-Automated and Reproducible Biological-Based Method to Quantify Calcium Deposition In Vitro
Published on: June 2, 2022
Development of a new measurement method for serum calcium with chlorophosphonazo-III
Eisaku Hokazono1, Susumu Osawa, Tomota Nakano
1Department of Health Sciences, Division of Biological Science and Technology, Graduate School of Medical Sciences, Kyushu University, 3-1-1 Maidashi, Higashi-ku, Fukuoka 812-8582, Japan. hokazono@shs.kyushu-u.ac.jp
Background:
Although serum calcium has been measured using the o-cresolphthalein complexone (oCPC) method in the clinical laboratory, this method still has some problems regarding linearity and reagent stability. We developed a new measurement procedure using chlorophosphonazo-III (CPZ-III: 2,7-bis (4-chloro-2-phosphonophenylazo) -1,8- dihydroxy-3, 6-naphthalenedisulphonic acid, disodium salt) as a chelator with an acid medium for serum calcium measurement. The present method showed better linearity and reagent stability compared with the oCPC method.
Methods:
Characteristics were studied in optimized conditions measuring wavelength by absorption spectra analysis, and interference of protein and metals with Mg(2+), Fe(2+), Cu(2+) and Zn(2+). The method was applied to an automated analyser (7170; Hitachi High Technologies Corp). The measurement performance was evaluated for accuracy, precision, recovery rate, linearity and reagent stability with a comparison study against atomic absorption spectrophotometry (AAS).
Results:
The within-run and between-run variations (coefficient of variation [CV]) were 0.92-1.01% and 0.75-1.43%, respectively. The linearity was 0-7.0 mmol/L. The comparison study obtained y = 1.002x (AAS) - 0.10, Sy/x = 0.18 mmol/L, n = 50. Reagent stability was at least 20 d at 4 degrees C without daily calibration.
Conclusion:
The new calcium measurement method in serum was demonstrated to have reliable and acceptable performances as a routine test in clinical laboratory.
