Abstract
The improving efficacy of many biological therapeutics and identification of low-level biomarkers are driving the analytical proteomics community to deal with extremely high levels of sample complexity relative to their analytes. Many protein quantitation and biomarker validation procedures utilize an immunoaffinity enrichment step to purify the sample and maximize the sensitivity of the corresponding liquid chromatography tandem mass spectrometry measurements. In order to generate surrogate peptides with better mass spectrometric properties, protein enrichment is followed by a proteolytic cleavage step. This is often a time-consuming multistep process. Presented here is a workflow which enables rapid protein enrichment and proteolytic cleavage to be performed in a single, easy-to-use reactor. Using this strategy Klotho, a low-abundance biomarker found in plasma, can be accurately quantitated using a protocol that takes under 5 h from start to finish.
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Conflict of Interest
Perfinity Biosciences is the manufacturer of the SMART Digest Immunoaffinity kit for Thermo Fisher Scientific.
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O’Grady, J.P., Meyer, K.W., Poe, D.N. (2017). Affinity Proteomics for Fast, Sensitive, Quantitative Analysis of Proteins in Plasma. In: Greening, D., Simpson, R. (eds) Serum/Plasma Proteomics. Methods in Molecular Biology, vol 1619. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-7057-5_5
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DOI: https://doi.org/10.1007/978-1-4939-7057-5_5
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