The cleavage pattern of F.IX and F.IXa treated with HNE as determined by reducing SDS-PAGE. Purified F.IX or F.IXa (4 μmol/L of each) was incubated with HNE (400 nmol/L) as described in the legend to Fig 1. Aliquots (4 μg protein) of F.IX (A) or F.IXa (B) were removed into reducing SDS sample after 0, 0.5, 2, 10, 20, and 30 minutes of incubation at 37°C with HNE (lanes 1 through 6, respectively), heated for 2 minutes at 90°C, electrophoresed in a 5% to 15% linear polyacrylamide gradient SDS gel according to Neville,20 and stained with Coomassie Blue. F.IX and F.IXa (A, lanes 7 and 8, respectively) or F.IXa (B, lane 7) incubated under the conditions given above for 30 minutes at 37°C but in the absence of added HNE is also shown. The positions of the molecular weight markers (in kilodaltons) are indicated to the left of each panel.