Fig. 6.

Analysis of the thrombin generation potential of microvesicle fractions separated by density gradient centrifugation. Samples of plasma from cancer patients (n=3) or pooled healthy plasma were cleared of any cell debris by centrifuging at 5,400g for 10 min on a microcentrifuge, and microvesicles were then sedimented at 20,000g and 100,000g at 20°C for 1 h. Microvesicles were then analysed by density gradient ultracentrifugation using a sucrose–OptiPrep gradient covering an approximate range of 1.02–1.22 g/ml against 2 sets of DensityMarkerBeads as density markers. The samples and markers were centrifuged at 52,000g for 90 min at 20°C. Following centrifugation, aliquots (0.5 ml) were sequentially removed. Thrombin generation activity in the fractions from cancer patients (A) and healthy plasma (B) was measured in the presence (circle) and absence of factor VII (square). (C) To highlight the TF-dependent activity, the average of the differences between thrombin generation in the presence and absence of factor VII was calculated (n=3).