Inhibition of thrombin generation by heparin and LMW heparins: a comparison of chromogenic and clotting methods.

Inhibition of thrombin generation by heparin and low-molecular-weight (LMW) heparins is an important parameter which relates to their anticoagulant actions in vivo. Previous studies in our laboratory used a clotting assay for assessment of thrombin generation but other published studies have used a chromogenic method. We have therefore measured the inhibition of thrombin generation by unfractionated heparin (UFH) and LMW heparins by a modified chromogenic method using microtitre plates and compared the results with the clotting method. The degree of inhibition of thrombin generation when calculated from both peak thrombin concentrations and areas under the curve in the chromogenic assay was the same. When the activities of each heparin were expressed as EC80, i.e. concentrations required for 80% inhibition of thrombin generation, the EC80 were higher in the chromogenic assay than in the clotting system. However when the potencies of the LMW heparins were expressed as percentages of that of the UFH standard by parallel line analysis, the differences between clotting and chromogenic assay results were small and not statistically significant (P > 0.05). This study demonstrates the feasibility of measuring inhibition of thrombin generation by a modified chromogenic method using microtitre plates, and shows that the results with this method are similar to those obtained with the clotting method.