Kenneth G. Mann,* Michael E. Nesheim, and Paula B. Tracy abstract: The molecular size and immunochemical properties of the unfractionatedfactor V present in plasma collected by venipuncture into a broad-spectrum anticoagulant and plate-let-inhibited mixture were compared with those reported for the isolated, single-chain factor V molecule of 330000 daltons. The anticoagulant-plasma mixture included 0.28% trisodium citrate, 1 mM benzamidine hydrochloride, 0.02% soybean trypsin inhibitor, 2.0 mM diisopropyl phosphorofluoridate, 10 mM dansylarginine 7V-(3-ethyl-1, 5-pentanediyl) amide, and 5 nM prostaglandin Ej. The Stokes radius of unfractionated factor V present in highly inhibited plasma (93 A) is virtually identical with the Stokes radius preidicted from the hydrodynamic data for the highly asymmetric, single-chain factor V molecule (91 A). With an expression which relates the Stokes radius and the sedimentation coefficient to the molecular weight of hydrodynamic units, the molecular weight obtained

Factor V was discovered by Owren (1947a), who, in 1943, identified a patient with a congenital lack of this coagulation factor. Subsequent work by Owren (1947b), Ware & Seegers (1948), and Murphy & Seegers (1948) identified factor V as one of the essential components required for the rapid con-version of prothrombin to thrombin. Early workers applied the term “labile factor” in discussing factor V because of the extreme instability of the protein during manipulation and storage. Because of this instability problem and the “procofactor” nature of the factor V (Nesheim et al., 1979a), the protein proved to be difficult to isolate. The first practical preparations of this protein were provided by Esnouf & Jobin (1967) and Barton & Hanahan (1967), and studies of preparations at this level of purity provided a great deal of insight into the function of factor V and the prothrombinase complex. Numerous subsequent re-ports of factor V purification have appeared in the literature (Colman, 1969; Dombrose & Seegers, 1974; Chulkova & Hernandez, 1975; Day, 1975; Smith & Hanahan, 1976; and Saraswathi et al., 1978). However, most preparations are complicated by the lack of homogeneity that is evident on electrophoretic analysis of the purified material and numerous conflicting assertions with regard to physical properties, sub-units, etc. There is little doubt that one of the principal difficulties confronting investigators attempting to isolate factor V is in the susceptibility of the protein to proteolysis during activation (Nesheim & Mann, 1979; Esmon, 1979) and inactivation (Canfield et al., 1978). Recently, two laboratories (Nesheim et al., 1978, 1979b; Esmon, 1979) have described preparation methods for factor V which provide a protein which is apparently homogeneous