ABSTRACT
Treatment with solvent/detergent is a widely used method for ensuring the virus safety of plasma products. In the present study, virus inactivation by a novel solvent/detergent combination, i.e. TnBP (tri-n-butyl phosphate) and polysorbate 20 during the manufacture of the factor VIII/VWF concentrate Optivate has been investigated. The inactivation of most enveloped viruses was rapid, i.e. > 5 log in 2 min, although the inactivation of vaccinia virus was slower, i.e. 4 log in 1h. Virus inactivation was effective over a wide range of conditions, i.e. solvent/detergent concentration, protein concentration and temperature, irrespective of whether tested individually or in combination. This confirms the effectiveness and robustness of this alternative version of the solvent/detergent procedure, and allows appropriate control limits to be set for this manufacturing step. Polysorbate 20 provides an alternative to the non-ionic detergents currently in use with the solvent/detergent procedure.
Subject(s)
Detergents/pharmacology , Factor VIII/standards , Polysorbates/pharmacology , Virus Inactivation , von Willebrand Factor/standards , Animals , Cell Survival/drug effects , Cells, Cultured , Chemical Precipitation , Chlorocebus aethiops , Drug Combinations , Drug Contamination/prevention & control , Factor VIII/chemical synthesis , Factor VIII/chemistry , Organophosphates/pharmacology , Solvents/pharmacology , Temperature , Vero Cells , Virus Diseases/blood , Virus Diseases/pathology , Virus Diseases/prevention & control , von Willebrand Factor/chemical synthesis , von Willebrand Factor/chemistryABSTRACT
We have studied the interaction of ASvWf with human platelets in PRP and in suspensions of washed platelets containing either physiological or low external ionized calcium concentration [Ca2+]o. In hirudin-PRP or in washed platelets in 1.5-2 mM CaCl2, ASvWf up to 50 micrograms/ml does not induce platelet aggregation or the release reaction. When [Ca2+]o is decreased by addition of citrate to hirudin-PRP or when no CaCl2 is added to washed platelet suspensions, ASvWf does induce platelet aggregation and the release reaction. In low [Ca2+]o, ASvWf interacts with platelet GPIb to cause primary aggregation of disc-shaped platelets to each other through GPIIb/IIIa, with or without added fibrinogen. This primary platelet aggregation leads to thromboxane A2 formation and secondary aggregation and the release reaction. With [Ca2+]o in the physiological range, there is less ASvWf interaction with GPIb, no primary platelet aggregation and no thromboxane A2 formation. The ASvWf-platelet interaction at physiological [Ca2+]o, however, enhances the platelet response to collagen or epinephrine.
