Methods used for monitoring coumarol treatment should be sensitive to the loss of calcium binding γ-carboxyl glutamic acids in the vitamin K dependent clotting factors. A two stage method sensitive to factors II-VII-X using the chromogenic substrate H-D-Phe-Pip-Arg-pNA has been evaluated using 75μl of reconstituted factor III containing 0.012μg of apoprotein III per ml for each assay. The reconstituted factor III used as the activator was prepared according to the method of Bjorklid et al. (1973) from human brain, human placental material and bovine brain, and from a phospholipid fraction - Gierksky et al. (1976).This method was sensitive to the coumarol induced defect with all three sources of apoprotein III and correlated well with a standard saline thrombo plastin extract (r=0.85, 0.89 and 0.88 respectively) in a group of 80 dicouroarol treated patients and 20 normal subjects and appears to overcome the insensitivity usually associated with crude bovine thromboplastins. Bjorklid E, Storm E and Prydz H (1973)Biochemical and Biophysical Research Communication, 55, 969
Gierksky K E, Bjorklid E and Prydz H (1976) Scandinavian Journal of Haematology,16, 300.