Can the diagnostic reliability of the thrombin generation test as a global haemostasis assay be improved? The impact of calcium chloride concentration.

BACKGROUND

Thrombin generation test (TGT) is a global haemostasis assay with a potential to predict bleeding tendencies and treatment effects in patients with haemophilia. Despite 15 years of clinical research, the diagnostic value of TGT remains controversial, possibly due to suboptimal sensitivity to coagulation deficiencies, robustness and reproducibility.

OBJECTIVE

The goal of this study was to explore the effect of calcium chloride (CaCl ) concentration on the TGT's response to intrinsic coagulation factors (F) VIII, IX and XIa.

METHODS

Normal and factor-deficient plasmas supplemented with lacking coagulation factor and different CaCl levels were tested by calibrated thrombinography assay.

RESULTS

Thrombin peak height (TPH) was strongly CaCl dependent, increasing sharply from no TG at 5 mm to a peak at 13.8 mm of CaCl (95% confidence interval [CI]: 13.0, 14.5) in normal and normalized deficient plasmas and at 11.9 mm (CI: 9.7, 14.2) in deficient plasmas, and then decreasing slowly to a complete inhibition at 30-40 mm. In contrast, TG lag time, time to peak and endogenous thrombin potential were nearly insensitive to CaCl concentrations between 10 and 20 mm. The maximal difference between the TPH in deficient and supplemented plasmas was observed at 15.5 mm (CI: 12.8, 18.1).

CONCLUSION

Variations in CaCl concentration in the assay mixture and sodium citrate concentrations in patient plasma samples may affect TGT responses, sensitivity and result in increased inter- and intra-laboratory variance. Implementation of TGT by clinical and quality control laboratories may require optimization of CaCl concentration.