Human Thrombin Production from Single Donor Plasma Using Thrombin Processing Device -TPD^™ .

Introduction: Thrombin is routinely combined in surgical practice with a fibrinogen source to prepare fibrin sealant to promote hemostasis or with platelet concentrates to prepare gels to enhance wound healing. Currently, thrombin is derived primarily from bovine tissue. The purpose of this study was to evaluate the robustness and reproducibility of the TPD, a new handheld sterile disposable designed to generate active human thrombin in less than 30 minutes from autologous or single donor allogeneic plasma. Thrombin is generated in the device from plasma via the activation of the alternative pathway of the coagulation cascade and thrombin inhibitors (e.g., antithrombin III) are concurrently depleted by a modified Cohn fractionation process. By eliminating the risk of the potential for allergic reactions and blood borne pathogen transmission, autologous human thrombin can be considered the safest thrombin for the preparation of fibrin sealant and platelet gels.

Materials and Methods: Active thrombin was prepared by combining 4 ml of thrombin reagent (a mixture of calcium chloride, ethanol and water) and 11 ml of plasma in a reaction chamber containing negatively charged particles (TPD). The mixture was incubated for 25 minutes and then the active thrombin serum harvested. The thrombin was prepared at ambient room temperatures of 18°C or 25°C (n=100/group) to assess robustness.

Results: The thrombin activity (mean ± S.D.) produced at 18°C and 25°C ambient temperature was 101 ± 22 IU/ml (n=100) and 90 ± 26 IU/ml (n=100), respectively. The average volume of thrombin serum harvested from the TPD was 9.2 ml in both groups. The thrombin concentration generated was demonstrated to rapidly (<5 seconds) coagulate fibrinogen concentrates at a 1:1 ratio (volume to volume). The thrombin maintained clotting activity for ≥4 hours storage at room temperature using a tilt clot test with cryoprecipitate as the source of fibrinogen (less than 5 second clot time).

Conclusions: The results demonstrate that the TPD is able to consistently generate high thrombin activity that is stable for ≥ 4 hours. There was no significant difference in thrombin production at the two extremes of the normal ambient temperature (18° C and 25° C) demonstrating the robustness of the device. The device offers a simple and rapid method for preparing active thrombin from autologous or single donor allogeneic human plasma so as to avoid the safety issues associated with bovine thrombin.