title: Tau Protein Quantification in Human Cerebrospinal Fluid by Targeted Mass Spectrometry at High Sequence Coverage Provides Insights into Its Primary Structure Heterogeneity.
abstract: Tau protein plays a major role in neurodegenerative disorders, appears to be a central biomarker of neuronal injury in cerebrospinal fluid (CSF), and is a promising target for Alzheimer's disease immunotherapies. To quantify tau at high sensitivity and gain insights into its naturally occurring structural variations in human CSF, we coupled absolute quantification using protein standard with the multiplex detection capability of targeted high-resolution mass spectrometry (MS) on a Quadrupole-Orbitrap instrument. Using recombinant tau we developed a step-by-step workflow optimization including an extraction protocol that avoided affinity reagents and achieved the monitoring of 22 tau peptides uniformly distributed along the tau sequence. The lower limits of quantification ranged (LLOQ) from 150 to 1500 pg/mL depending on the peptide. Applied to endogenous CSF tau, up to 19 peptides were detected. Interestingly, there were significant differences in the abundance of peptides depending on their position in the sequence, with peptides from the tau mid-domain appearing significantly more abundant than peptides from the N- and C-terminus domains. This MS-based strategy provided results complementary to those of previous ELISA or Western Blot studies of CSF tau and could be applied to tau monitoring in human CSF cohorts.
authors.affiliation: CEA, iBiTec-S , Service de Pharmacologie et d'Immunoanalyse, Laboratoire d'Etude du Métabolisme des Médicaments, Gif-sur-Yvette 91191, France.|CHU Montpellier, IRMB, hôpital St Eloi, Laboratoire de Biochimie Protéomique Clinique et CCBHM , INSERM-UM1 U1040, Montpellier F-34000 France.
authors.forename: Nicolas R
authors.initials: NR
authors.lastname: Barthélemy
authors.affiliation: CEA, iBiTec-S , Service de Pharmacologie et d'Immunoanalyse, Laboratoire d'Etude du Métabolisme des Médicaments, Gif-sur-Yvette 91191, France.
authors.forename: François
authors.initials: F
authors.lastname: Fenaille
authors.affiliation: CHU Montpellier, IRMB, hôpital St Eloi, Laboratoire de Biochimie Protéomique Clinique et CCBHM , INSERM-UM1 U1040, Montpellier F-34000 France.
authors.forename: Christophe
authors.initials: C
authors.lastname: Hirtz
authors.affiliation: Inserm, UMR-S 1172, Alzheimer & Tauopathies, Centre de Recherche Jean-Pierre Aubert, Lille, Univ. Lille, Faculté de Médecine, Lille F-59045, France.
authors.forename: Nicolas
authors.initials: N
authors.lastname: Sergeant
authors.affiliation: Inserm, UMR-S 1172, Alzheimer & Tauopathies, Centre de Recherche Jean-Pierre Aubert, Lille, Univ. Lille, Faculté de Médecine, Lille F-59045, France.
authors.forename: Susanna
authors.initials: S
authors.lastname: Schraen-Maschke
authors.affiliation: CHU Montpellier, IRMB, hôpital St Eloi, Laboratoire de Biochimie Protéomique Clinique et CCBHM , INSERM-UM1 U1040, Montpellier F-34000 France.
authors.forename: Jérôme
authors.initials: J
authors.lastname: Vialaret
authors.affiliation: Inserm, UMR-S 1172, Alzheimer & Tauopathies, Centre de Recherche Jean-Pierre Aubert, Lille, Univ. Lille, Faculté de Médecine, Lille F-59045, France.
authors.forename: Luc
authors.initials: L
authors.lastname: Buée
authors.affiliation: CHU Montpellier, IRMB, hôpital St Eloi, Laboratoire de Biochimie Protéomique Clinique et CCBHM , INSERM-UM1 U1040, Montpellier F-34000 France.|Centre Mémoire Ressources Recherche, CHU Montpellier, hôpital Gui de Chauliac, Montpellier. Université Montpellier I , Montpellier F-34000 France.
authors.forename: Audrey
authors.initials: A
authors.lastname: Gabelle
authors.affiliation: CEA, iBiTec-S , Service de Pharmacologie et d'Immunoanalyse, Laboratoire d'Etude du Métabolisme des Médicaments, Gif-sur-Yvette 91191, France.
authors.forename: Christophe
authors.initials: C
authors.lastname: Junot
authors.affiliation: CHU Montpellier, IRMB, hôpital St Eloi, Laboratoire de Biochimie Protéomique Clinique et CCBHM , INSERM-UM1 U1040, Montpellier F-34000 France.
authors.forename: Sylvain
authors.initials: S
authors.lastname: Lehmann
authors.affiliation: CEA, iBiTec-S , Service de Pharmacologie et d'Immunoanalyse, Laboratoire d'Etude du Métabolisme des Médicaments, Gif-sur-Yvette 91191, France.
authors.forename: François
authors.initials: F
authors.lastname: Becher
mesh_terms: D000595:Amino Acid Sequence; D015415:Biomarkers / Q000134:cerebrospinal fluid; D002853:Chromatography, Liquid; D006801:Humans; D013058:Mass Spectrometry / Q000295:instrumentation* / Q000379:methods*; D008969:Molecular Sequence Data; D010455:Peptides / Q000134:cerebrospinal fluid* / Q000737:chemistry; D015203:Reproducibility of Results; D016875:tau Proteins / Q000134:cerebrospinal fluid* / Q000737:chemistry
chemical_list: D015415:Biomarkers; D010455:Peptides; D016875:tau Proteins
keywords: alternative splicing-dependent peptides; cerebrospinal fluid; microtubule-associated tau protein; parallel reaction monitoring; protein absolute quantification; protein fragments; protein precipitation; solid-phase extraction