Coagulation Factor VIII is a heavily glycosylated heterodimeric plasma protein that consists of a heavy (domains A1-A2-B) and light chain (domains A3-C1-C2). Factor VIII participates in the intrinsic pathway of blood coagulation. Defects in the Factor VIII gene result in haemophilia A. Both variants, plasma-derived and recombinant Factor VIII products are available on the market. With its high degree of complex glycosylation (25 N-glycosylation sites and 6 O-glycosylation sites) and a size of ca. 265 kDa (2351 amino acids) this molecule is very challenging from an analytical point of view.

To support the development of Factor VIII biosimilars or biobetters, Protagen Protein Services has set up an analysis program in accordance with ICH Q6B guidelines, including method setup, validation of customer-specific methods, and release testing.

This package includes protein quantification, characterizing the protein primary structure, analysis of N- glycosylation and protein modifications. The analysis also allows to monitor the Tyrosine-sulfation levels, which are important for the functionality of Factor VIII.

The following analytical methods can be provided for Factor VIII:
  • Quantity and Extinction Coefficient:
    • Amino acid analysis: determination of protein content and amino acid composition
    • Determination of extinction coefficient
  • Structural Characterization and Confirmation – Amino Acid Sequence:
    • Peptide mapping
    • Deamidation, methionine oxidation and tyrosine sulfation levels
    • Disulfide bridge analysis by mass spectrometry
    • N/C terminal sequencing: MALDI-ISD
    • Total molecular weight of each subunit
  • Structural Characterization and Confirmation – Glycosylation:
    • Monosaccharide composition
    • Sialic acid content
    • N- and O-Glycan mapping (MALDI MS) and Linkage analysis by GC-MS
    • N- and O-Glycosylation site determination
  • Physicochemical Properties:
    • Isoform pattern (IEF)
    • Liquid chromatographic pattern: IEX
    • Spectroscopic profiles: UV/VIS Spectra
    • Spectroscopic profiles: CD
    • SEC-MALLS for aggregation

Your contact for Factor VIII at Protagen Protein Services GmbH

Martin-Blueggel 150x100px

Martin Blüggel

Chief Executive Officer (CEO)
Our business development team will be happy to assist you with your project.

Phone: +49 (0) 231 9742-6100