Abstract
Coagulation factor VIII (FVIII) is an important glycoprotein involved in the extrinsic coagulation cascade. Mutations in FVIII gene results in hemophilia A, a recessive coagulation disorder that is clinically managed by administration of purified FVIII from blood donors or recombinant FVIII. Because of its fundamental therapeutic application, biotechnological production of FVIII requires rigid quality control and monitoring in patients and clinical trials. Here, we describe a protocol for a mass spectrometry based approach termed selective reaction monitoring (SRM) as an important alternative tool for accurate and sensitive quantitation of purified or recombinant FVIII.
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References
CA SB, Lim A (2007) Applications of mass spectrometry for the structural characterization of recombinant protein pharmaceuticals. Mass Spectrom 26:370–388
Picotti P, Aebersold R (2012) Selected reaction monitoring-based proteomics: workflows, potential, pitfalls and future directions. Nat Methods 9:555–566
Liebler D, Zimmerman LJ (2013) Targeted quantification of proteins by mass spectrometry. Biochemistry 52:3797–3806
Kuster B, Schirle M, Mallick P, Aebersold R (2005) Scoring proteomes with proteotypic peptide probes. Nat Rev Mol Cell Biol 6(7):577–583
Deutsch EW, Lam H, Aebersold R (2008) PeptideAtlas: a resource for target selection for emerging targeted proteomics workflows. EMBO Rep 9:429–434
Maclean B, Tomazela DM, Shulman N, Chambers M, Finney GL, Frewen B, Kern R, Tabb DL, Liebler DC, Maccoss MJ (2010) Skyline: an open source document editor for creating and analyzing targeted proteomics experiments. Bioinformatics 26:966–968
Russo-Carbolante EMS, Swiech K, Covas DT, Picanço-Castro V (2015) Large scale and stable production of human FVIII in the human cell line Sk-Hep-1. US Patent 8,969,041 B2
Rosa NG, Swiech K, Picanço-castro V, Russo-carbolante EMS, Neto MSA, Fernandes AC, Faça VM, Covas DT (2012) SK-HEP cells and lentiviral vector for production of human recombinant factor VIII. Biotechnol Lett 34(8):1435–1443
Apweiler R, Bairoch A, Wu CH, Barker WC, Boeckmann B, Ferro S, Gasteiger E, Huang H, Lopez R, Magrane M, Martin MJ, Natale DA, O’donavan C, Redashi N, Yeh LSL (2004) UniProt: the universal protein knowledgebase. Nucleic Acids Res 32:115–119
Acknowledgments
This research was supported by FAPESP (Young Scientist Grants—Proc. No. 2011/09347-1), Center for Cell Based Thereapy—CTC-CEPID (Proc. FAPESP 2013/08135-2) and CISBi-NAP. G.P.L. received a fellowship from CNPq Proc. No. 130142/2015-8, and V.M.F. received a fellowship from CNPq, Proc. No. 308561/2014-7.
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do Nascimento Filho, E.G., Lanfredi, G.P., Abreu-Neto, M.S., Covas, D.T., Faça, V.M. (2018). Quantification of Coagulation Factor VIII by Selective Reaction Monitoring. In: Picanço-Castro, V., Swiech, K. (eds) Recombinant Glycoprotein Production. Methods in Molecular Biology, vol 1674. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-7312-5_22
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DOI: https://doi.org/10.1007/978-1-4939-7312-5_22
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Publisher Name: Humana Press, New York, NY
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Online ISBN: 978-1-4939-7312-5
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