Performance of a recombinant fusion protein linking coagulation factor IX with recombinant albumin in one-stage clotting assays.

BACKGROUND: Measuring factor IX activity (FIX:C) with one-stage clotting (OSC) assays, based on the activated partial thromboplastin time (aPTT), is the current mainstay of diagnostic techniques for hemophilia B. Assessing the performance of new recombinant FIX (rFIX) products in OSC assays is essential as aPTT reagents from different manufacturers yield different potency estimates for rFIX.

OBJECTIVES: To evaluate the extent to which choice of reagent composition influences rFIX potency measurements of recombinant FIX-albumin fusion protein (rIX-FP, IDELVION® ) activity in OSC assays.

METHODS: rIX-FP was added to FIX-deficient plasma and FIX:C assessed centrally and locally in a multicenter international field study with a variety of commercial OSC aPTT reagents. Paired sample analysis of clinical samples compared values of FIX:C from local and central laboratories. In-house bioanalytic investigations with spiked samples were conducted to compare the aPTT-reagent dependent variability of rIX-FP with unmodified rFIX and rFIX Fc fusion protein (rFIXFc).

RESULTS: Central and local assessments of FIX:C from 10 countries and 21 participating centers showed comparable results to the central laboratory across the majority of 18 different aPTT reagents from both clinical and spiked samples. There was a consistent underestimation in rIX-FP activity of approximately 50% with OSC assays using Actin® FS or kaolin-based aPTT reagents. In the bioanalytical study, rIX-FP showed comparable variability in OSC assays to unmodified rFIX and rFIXFc.

CONCLUSIONS: rIX-FP activity can be accurately measured using OSC assays with the majority of commercial reagents. Actin® FS or kaolin-based reagents will likely lead to 50% underestimation in activity. This article is protected by copyright. All rights reserved.