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. Author manuscript; available in PMC: 2023 Mar 1.
Published in final edited form as: J Thromb Haemost. 2021 Dec 17;20(3):574–588. doi: 10.1111/jth.15616

Figure 2. Biochemical characterization of FVIII variants.

Figure 2.

FVIII activity of expressed FVIII supernatant by transfected BHK-M cells at ~70% confluency at ~1 × 106 BHK-M cells for BDD FVIII and FVIII variants determined by the aPTT-based one-stage clotting assay at 24 hours (A) and 48 hours (B). FVIII activity (A and B) are presented as means ± SD of 2-5 measurements per FVIII variant. BDD FVIII activity measurements reflect 4 representative and separate preparations of the protein and the modified FVIII variant reflect one preparation of the proteins. SDS-PAGE of BDD FVIII (C) and FVIII variants (D) are shown. SDS-PAGE was performed in the presence and absence of human thrombin (IIa). A1, FVIII A1 domain; A2, FVIII A2 domain; A1/A2, non-covalently linked A1/A2 domains; LC, FVIII light chain comprised of the A3, C1, and C2 domains; SC, FVIII single chain comprised of the A1, A2, A3, C1, and C2 domains. Binding of BDD FVIII and FVIII variants to human-derived VWF by ELISA is shown (E). Differences in the mean FVIII activity of each of the FVIII variants were compared to the mean activity of BDD FVIII by one-way ANOVA with Dunnett’s correction for multiple comparison. *P <0.05 and ***P <0.001.