Inter-laboratory comparison of plasma efanesoctocog alfa measurements

Introduction Efanesoctocog alfa (efa) is a recombinant coagulation factor VIII (FVIII) concentrate, engineered for improved extended half-life in hemophilia A treatment. Its design results in discrepancies in FVIII diagnostic assays. An interlaboratory study was conducted by the GTH-STAEKOLA to compare assay results as determined by various one-stage clotting (OSCA) and chromogenic substrate assays (CSA). Method Twenty study sample kits each containing 10 samples including efa spiked to congenital FVIII-deficient plasma (5 to 80 IU/dL) as well as post-infusion samples obtained at peak (30 min post-infusion) and trough levels (after 3.5 or 7 days) were prepared and shipped to 20 participating laboratories (17x D, 2x A, and 1x CH). Analysis was done using various OSCAs / aPTT-reagents (Actin FSL [n=10], Actin FS [9], Pathromtin SL [10], SynthASil [5], SynthAFax [1], aPTT-SP [1], CK Prest [1], and Roche FVIII [2]) and/or CSAs (Siemens FVIII [10], BIOPHEN FVIII [6], and Electrachrome FVIII [2]) at routine calibrations. Regarding assessment of results obtained for the post-infusion samples, FVIII activities obtained by Actin FSL-based OSCA-analysis were deemed true values. Results Analysis of spiked samples revealed acceptable results (+/- 30% of input value) for Actin FSL-, Pathrombin SL-, and CK Prest-based OSCAs at levels>20 IU/dL. In contrast, underestimation was observed for SynthASil-based OSCA at levels>20 IU/dL while overestimation by a factor of approx. 2 was found for Actin FS-based and the Roche FVIII OSCA. As previously described, overestimation by a factor of 2 to 3 was found when applying FVIII CSAs. Within the class of OSCAs, analysis of included post-infusion samples yielded expected deviations between assays. When analyzed by FVIII CSA, however, samples at trough levels yielded lower values than would have been expected based on spiked sample analysis. This corresponds to the experience of several laboratories, which have observed that chromogenic test results in the low activity range approximate those obtained by Actin FSL-based OSCA. Conclusion Actin FSL-, Pathrombin SL-, and CK Prest-based OSCA appear to be the methods of choice for measurement of efa in post-infusion samples. However, overestimation by a factor of approx. 1.5 must be considered at efa plasma levels of12 IU/dL) or the Roche FVIII OSCA (x0.6). Application of a FVIII CSA based on bovine factors may be indicated for determination of efa-activity in presence of emicizumab. Based on the observations described here, application of the generic CF of /2.5 described in the IFU is not recommended at trough levels. In general, proposed product-specific calibration appears to be a reasonable approach. Publication History Article published online: 17 February 2026 © 2026. Thieme. All rights reserved. Georg Thieme Verlag KG Oswald-Hesse-Straße 50, 70469 Stuttgart, Germany