Summary
The products of the reaction of activated factor IX with factor VIII, phospholipid
and calcium have been studied using gel filtration on Sephadex G-200. Both factor
VIII and phospholipid are completely excluded from the gel (distribution coefficient
or KD of zero) while factor IXa behaves like a protein similar in size to albumin (KD approx. 0.4). Factor IXa complexes with phospholipid when calcium ions are added to the eluting buffer. When
factor VIII, factor IXa, phospholipid and calcium are applied to the column and eluted with added Ca++ in the eluting buffer, a labile product with a KD of 0 is formed. The product apparently
dissociates in the absence of added calcium ions in the eluting buffer. In the presence
of calcium ions, the distribution coefficient of factor IX, when applied alone to
the column, increased. The above findings are analogous to those reported between
activated factor X, thrombin-activated factor V, phospholipid and calcium occurring
later in the coagulation process.