The use of the new ReFacto AF Laboratory Standard allows reliable measurement of FVIII:C levels in ReFacto AF mock plasma samples by a one‐stage clotting assay

Summary. Factor VIII coagulant (FVIII:C) levels measured in patients receiving ReFacto^® (B‐domain‐deleted recombinant FVIII) using chromogenic substrate assay (CSA) and one‐stage clotting assay (OSA) have frequently shown discrepancies, and the use of the ReFacto Laboratory Standard (RLS) has therefore been recommended to minimize these differences. The potency of ReFacto AF^®, the albumin‐free successor of ReFacto^®, is determined using CSA for the titration of vials, and a new standard (RLS‐AF) was developed to measure its biological efficacy using OSA. This multicentre study therefore evaluated the efficacy of this new RLS in minimizing differences between OSA and CSA when measuring FVIII:C levels in plasma. Mock plasma samples were prepared by diluting ReFacto AF^® in FVIII‐deficient plasma to obtain four concentrations ranging from 15 to 90 IU dL^?1. FVIII:C levels were then measured in six laboratories on four separate days using three different procedures, i.e. OSA with a plasma standard (PS) as reference, OSA with RLS‐AF and CSA with PS. The inter‐centre standard deviation ranged from 1.4 to 5.5 IU dL^?1. However, FVIII:C levels measured with OSA were closer to the expected values when RLS‐AF was used. In addition, the uncertainty of measurement, reflecting the inter‐method discrepancy was greatly reduced when RLS‐AF was employed in OSA (15%) in place of PS (33%). This study demonstrates that the OSA performed with RLS‐AF to establish calibration curves provides a valuable alternative to CSA to measure FVIII:C in ReFacto‐AF‐treated patients.