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Isocitrate as Calcium Ion Activity Buffer in Coagulation Assays

Division of Clinical Chemistry, Department of Biomedicine and Surgery, Linköping University, S-581 85 Linköping, Sweden.
^a Author for correspondence. Fax 46 13 223240; e-mail tony.gojceta{at}mbox200.swipnet.se

Background: Ca²⁺ activity close to the physiological concentration of 1.3 mmol/L is essential in blood coagulation. Is this also true for the performance of global diagnostic coagulation assays? We searched for compounds that would buffer Ca²⁺ activity at ~1.3 mmol/L without disturbing coagulation reactions and investigated whether such Ca²⁺ buffering improves diagnostic efficacy in global diagnostic coagulation tests.

Methods: Buffering was investigated by mixing CaCl₂ and 11 candidate compounds and determining Ca²⁺ activity. The best candidates were added to mixtures of plasma and thromboplastin to detect interference with coagulation reactions. The best of these candidates, isocitrate, was used to modify an activated partial thromboplastin time (APTT), buffering final Ca²⁺ activity to ~1.3 mmol/L. Plasma samples from 22 healthy individuals and 120 patients were analyzed with original and modified APTT to determine whether diagnostic efficacy was improved.

Results: Two suitable Ca²⁺ buffers, citrate and isocitrate, were found. Isocitrate was preferred as being less coagulation inhibitory, a better Ca²⁺ buffer, and possibly a better anticoagulant. The isocitrate-modified APTT showed a final Ca²⁺ activity of 1.60 ± 0.07 mmol/L, compared with 2.73 ± 0.20 mmol/L for the original APTT. The means and SDs for the healthy individuals were determined for both procedures, and the values were used to express patient deviation from normality (difference from mean divided by SD). The deviation was greater for the modified APTT; 4.3 ± 5.7, compared with 3.6 ± 5.0 (P <0.005) for the original APTT.

Conclusions: Isocitrate can be used to buffer Ca²⁺ activity at physiological concentrations and can serve as an anticoagulant. APTT with isocitrate-buffered Ca²⁺ activity shows signs of improved diagnostic efficacy.© 1999 American Association for Clinical Chemistry

The following articles in journals at HighWire Press have cited this article:

				J. Horsti Measurement of Prothrombin Time in EDTA Plasma with Combined Thromboplastin Reagent, Clin. Chem., November 1, 2000; 46(11): 1844 - 1846. [Full Text] [PDF]

				J. A. Koepke More on Calcium Ion Activity Buffers for Coagulation Testing Clin. Chem., September 1, 1999; 45(9): 1575 - 1576. [Full Text] [PDF]