For APTT investigation in plasma, fresh blood from healthy BABL/c mice was collected. Mice plasma was prepared by centrifuging citrated blood at 3000 × g for 10 min at 4 °C to remove blood cells. Fifty microliters of plasma was mixed with 50 µl TriniClot APTT reagent for 5 min incubation, followed by 16.7 µl Aptarray (~560 nM, containing 20 μM TBA15 and 20 μM HD22), T-16A-H (20 μM) or other control groups at 37 °C for 5 min. After the addition of 50 µl CaCl2 to initiate the coagulation, the clotting reaction of mice plasma was monitored by a semi-automatic coagulation analyzer (SC 40, STEELLEX).

For in vivo anticoagulation, BABL/c mice were administrated of buffer (100 μl), the mixture of origami and aptamers (Ori + T + H, aptamers 20 μM and origami ~560 nM), T-16A-H (20 μM) or Aptarray (~560 nM, containing 20 μM TBA15 and 20 μM HD22) via a single tail vein injection. Blood samples collected from mice were anticoagulated with 3.2% sodium citrate at 5 min. Whole mice blood was centrifuged at 3000 × g for 10 min at 4 °C to remove blood cells. The collected plasma sample was mixed with 50 µl TriniClot APTT regent for 5 min incubation. After the addition of 50 µl CaCl2 to the solution, the clotting reaction of mice plasma was monitored by a semi-automatic coagulation analyzer.

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