Isocitrate as calcium ion activity buffer in coagulation assays

Background: Ca2+ activity closets the physiological concentration of 1.3 mm ol/L is essential in blood coagulation, Is this also true for the performan ce of global diagnostic coagulation assays? We searched for compounds that would buffer Ca2+ activity at similar to 1.3 mmol/L without disturbing coag ulation reactions and investigated whether such Ca2+ buffering improves dia gnostic efficacy in global diagnostic coagulation tests. Methods: Buffering was investigated by mixing CaCl2 and II candidate compou nds and determining Ca2+ activity. The best candidates were added to mixtur es of plasma and thromboplastin to detect interference with coagulation rea ctions. The best of these candidates, isocitrate, was used to modify an act ivated partial thromboplastin time (APTT), buffering final Ca2+ activity to similar 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. Iso citrate was perferred as being less coagulation inhibitory, a better Ca2+ b uffer, and possibly a better anticoagulant, The isocitrate-modified APTT sh owed 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 indivi duals were determined for both procedures, and the values were used to expr ess 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 physiologica l concentrations and can serve as an anticoagulant. APTT with isocitrafe-bu ffered Ca2+ activity shows signs of improved diagnostic efficacy. (C) 1999 American Association for Clinical Chemistry.