Triniclot aptt reagent

Factor Xa inhibitors were supplemented in citrated plasma over a concentration range of 1.0 to 0.062 µg/mL. Either saline as a control or AA at a final concentration of 100 µg/mL was added to individual aliquots of plasma supplemented with each inhibitor. Samples were analyzed using the 1-stage prothrombinase-induced clotting time (PiCT), activated partial thromboplastin time (aPTT), and prothrombin time (PT) assays. The PiCT assay reagents were obtained from PentaPharm (Basel, Switzerland). The TriniCLOT aPTT reagent was obtained from Diagnostica Stago (Parsippany, New Jersey). The PT, HemosIL reagent was obtained from Instrumentation Laboratory (Bedford, Massachusetts). All reagents were reconstituted according to the manufacturer’s instructions. Activated-partial thromboplastin time and PT were measured using an ACL-Elite (Instrumentation Laboratory). Prothrombinase-induced clotting time was measured using an ST4 instrument from Diagnostica Stago (Parsippany, New Jersey). Results were compiled in terms of mean (SD).

Whole Blood Analysis: Hemochron whole blood coagulation system (Accriva Diagnostics, Inc. CA, USA) was used to measure whole blood activated clotting time (ACT). Whole blood was supplemented with each heparin at a final concentration of 2.5 µg/ml as control and transferred into a celite ACT tube. For reversal studies, blood was supplemented with the heparins at a final concentration of 2.5 µg/ml were either supplemented with AA at a final concentration of 100 µg/ml or PrSO₄ at a final concentration of 25 µg/ml and transferred into celite ACT tubes. Results were recorded in seconds and compiled in terms of means ± SD.
Plasma Supplemented System: Each of the heparins were supplemented in citrated plasma over a concentration range of 10 - 0.62 µg/ml. Saline as a control, AA at a final concentration of 100 µg/ml, and PrSO₄ at 10 µg/ml were added to individual aliquots of plasma supplemented with each drug. The TriniCLOT - aPTT reagent was obtained from Diagnostica Stago (Parsippany, NJ, USA) and used to analyze activated partial thromboplastin time (aPTT) by using an ACL-Elite (Instrumentation Laboratory, Bedford, MA, USA). Results were compiled in terms of mean   SD.

Each of the agents were supplemented in citrated plasma over a concentration
range of 0.0-10.0 µg/mL. Saline was used for referencing purpose. For
prothrombinase-induced clotting time (PiCT), reagents were obtained from
PentaPharm (Basel, Switzerland) and samples were analyzed by using 2-stage
technique. For activated partial thromboplastin time (aPTT) testing, the
TriniCLOT aPTT reagent was obtained from Diagnostica Stago (Parsippany, New
Jersey, USA). Alfa human thrombin was obtained for thrombin time (TT) from
Enzyme Research Laboratories (South Bend, Indiana, USA). The PT, HemosIL reagent
was obtained from Instrumentation Laboratory (Bedford, Massachusetts, USA).
Thrombin was reconstituted with 0.025 M CaCl₂ at a concentration of 5
U/ml which was used to measure TT. ACL-Elite (Instrumentation Laboratory,
Bedford, Massachusetts, USA) was used to analyze aPTT, TT and PT. The PiCT test
was measured on the ST4 clot analyzer (Diagnostica Stago, Paris, France).
Results were compiled in terms of average mean ± SD of 3 runs.