The Large Scale Preparation of Clottable Fibrinogen-Free, High Purity, High Potency Factor VIII Concentrate

Crude Factor VIII was initially extracted from plasma as cryoprecipitate. This crude concentrate was treated with high molecular weight polymer F-48 to remove the bulk of fibrinogen. The yield of Factor VIII at this point was 6356 theoretical and 42% actual (the starting plasma contained,66 units/ml VIII), Total protein in the cryoprecipitate was 2,2 gm# and 0.93 gn# after removal of fibrinogen bulk. The next step was addition of a thrombin-like enzyme, at a concentration of 0.5 units, with the resultant removal of remaining fibrinogen. After clotting out residual fibrinogen, resultant fibrin strands were removed by high speed centrifugation. The final product gave a theoretical yield of 65% and an actual yield of 28%; thus, 65% of the initial Factor VIII was recovered in the final product. Prior to lyophilization, the final product was stabilized with albumin. After lyophilization, no loss of activity occurred and solubility was excellent. Protein electrophoretic analysis revealed 70% in the albumin region, 7% in the alpha & beta globulin region, and 20% in the gamma region. Tests for hemolysins were negative and isoagglutin-in titers were 1:6. Plasminogen and plasmin were undetectable and fibrin(ogen) degradation products were 80 ug/ml. The large scale preparation of this fibrinogen-free Factor VIII concentrate may prove highly useful in sparing the hemophilic patient fibrin(ogen) deposits in the kidneys and other organs, a complication of existing concentrates. In addition, much higher potency in much less volume may be achieved with this material. The cost for large scale preparation of this material should be the same as for existing concentrates.