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J Proteomics. 2018 Mar 20;175:12-26. doi: 10.1016/j.jprot.2017.08.003. Epub 2017 Aug 30.

Towards a routine application of Top-Down approaches for label-free discovery workflows.

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Bruker Daltonique S.A., 34, rue de l'industrie, 67160 Wissembourg, France. Electronic address:
Laboratoire de Biochimie et Protéomique Clinique, Institut de Médecine Régénératrice et de Biothérapie, CHU de Montpellier - Hôpital St. Eloi, 34000 Montpellier, France.
Radboud Proteomics Center, Translational Metabolic Laboratory, Department of Laboratory Medicine, Radboud Center for Mitochondrial Medicine, Radboud University Medical Center, Geert Grooteplein Zuid 10, 6525 GA Nijmegen, The Netherlands.
Laboratoire de Biochimie et Protéomique Clinique, Institut de Médecine Régénératrice et de Biothérapie, CHU de Montpellier - Hôpital St. Eloi, 34000 Montpellier, France; Centre Mémoire Ressources Recherche, CHU Montpellier, hôpital Gui de Chauliac, Montpellier, Université Montpellier I, Montpellier, F-34000, France.
Bruker Daltonics Inc., 40 Manning Road, Billerica, MA 01821, USA.
Protein Metrics Inc., 1622 San Carlos Ave., San Carlos, CA 94070, USA.
Bruker Daltonik GmbH, Fahrenheitstrasse 4, 28359 Bremen, Germany.


Thanks to proteomics investigations, our vision of the role of different protein isoforms in the pathophysiology of diseases has largely evolved. The idea that protein biomarkers like tau, amyloid peptides, ApoE, cystatin, or neurogranin are represented in body fluids as single species is obviously over-simplified, as most proteins are present in different isoforms and subjected to numerous processing and post-translational modifications. Measuring the intact mass of proteins by MS has the advantage to provide information on the presence and relative amount of the different proteoforms. Such Top-Down approaches typically require a high degree of sample pre-fractionation to allow the MS system to deliver optimal performance in terms of dynamic range, mass accuracy and resolution. In clinical studies, however, the requirements for pre-analytical robustness and sample size large enough for statistical power restrict the routine use of a high degree of sample pre-fractionation. In this study, we have investigated the capacities of current-generation Ultra-High Resolution Q-Tof systems to deal with high complexity intact protein samples and have evaluated the approach on a cohort of patients suffering from neurodegenerative disease. Statistical analysis has shown that several proteoforms can be used to distinguish Alzheimer disease patients from patients suffering from other neurodegenerative disease.


Top-down approaches have an extremely high biological relevance, especially when it comes to biomarker discovery, but the necessary pre-fractionation constraints are not easily compatible with the robustness requirements and the size of clinical sample cohorts. We have demonstrated that intact protein profiling studies could be run on UHR-Q-ToF with limited pre-fractionation. The proteoforms that have been identified as candidate biomarkers in the-proof-of concept study are derived from proteins known to play a role in the pathophysiology process of Alzheimer disease.


Alzheimer disease; Clinical proteomics; Top-down label-free proteoform profiling; Ultra-high resolution Q-Tof

[Indexed for MEDLINE]

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