Abstract

Quantitative Mass Spectrometry of Peptides and Proteins, A New Era in Clinical Biomarker Detection

Sylvain Lehmann, Professor, University Hospital Montpellier

Recent improvements in mass spectrometry (MS) allow this technology to quantify with clinical grade analytical sensitivity and specificity, peptides and proteins in biological fluids. We believe that in some cases MS will represent a valuable alternative to traditional methods of immunochemical quantification of proteins. We followed this path for the quantitation of two analytes of interest. The first one is hepcidin, a small peptide involved in iron metabolism which immune-detection is very challenging (as the folded structure of this peptide results in a low immunogenicity, aggregability and eventually in a high variability). Diagnosis applications related to hepcidin measurement are numerous, and involve iron-overload disorders or anemia in inflammatory context (critical care, cancer..). The second one is the apolipoprotein E (ApoE) which has three isoforms (e2, e3 and e4) and is a major risk factor of Alzheimer and cardiovascular diseases. For this purpose, quantitative targeted mass spectrometry (SRM/MRM) was developed using a triple quadripole. Sample prefractionation, trypsic digestion and sample clean-up were realised using an automated liquid handling robot. Quantification relied on internal isotope labeled synthetic peptides as standard. In conclusion, fully validated LC-MS/MS methods were developed for the determination of hepcidin and ApoE providing interesting examples how MS could contribute to development and standardization of clinical chemistry assays.