title: Interlaboratory study on differential analysis of protein glycosylation by mass spectrometry: the ABRF glycoprotein research multi-institutional study 2012.
abstract: One of the principal goals of glycoprotein research is to correlate glycan structure and function. Such correlation is necessary in order for one to understand the mechanisms whereby glycoprotein structure elaborates the functions of myriad proteins. The accurate comparison of glycoforms and quantification of glycosites are essential steps in this direction. Mass spectrometry has emerged as a powerful analytical technique in the field of glycoprotein characterization. Its sensitivity, high dynamic range, and mass accuracy provide both quantitative and sequence/structural information. As part of the 2012 ABRF Glycoprotein Research Group study, we explored the use of mass spectrometry and ancillary methodologies to characterize the glycoforms of two sources of human prostate specific antigen (PSA). PSA is used as a tumor marker for prostate cancer, with increasing blood levels used to distinguish between normal and cancer states. The glycans on PSA are believed to be biantennary N-linked, and it has been observed that prostate cancer tissues and cell lines contain more antennae than their benign counterparts. Thus, the ability to quantify differences in glycosylation associated with cancer has the potential to positively impact the use of PSA as a biomarker. We studied standard peptide-based proteomics/glycomics methodologies, including LC-MS/MS for peptide/glycopeptide sequencing and label-free approaches for differential quantification. We performed an interlaboratory study to determine the ability of different laboratories to correctly characterize the differences between glycoforms from two different sources using mass spectrometry methods. We used clustering analysis and ancillary statistical data treatment on the data sets submitted by participating laboratories to obtain a consensus of the glycoforms and abundances. The results demonstrate the relative strengths and weaknesses of top-down glycoproteomics, bottom-up glycoproteomics, and glycomics methods.
authors.affiliation: Center for Biomedical Mass Spectrometry, Boston University School of Medicine, Boston, Massachusetts 02118;
authors.forename: Nancy
authors.initials: N
authors.lastname: Leymarie
authors.forename: Paula J
authors.initials: PJ
authors.lastname: Griffin
authors.forename: Karen
authors.initials: K
authors.lastname: Jonscher
authors.forename: Daniel
authors.initials: D
authors.lastname: Kolarich
authors.forename: Ron
authors.initials: R
authors.lastname: Orlando
authors.forename: Mark
authors.initials: M
authors.lastname: McComb
authors.forename: Joseph
authors.initials: J
authors.lastname: Zaia
authors.forename: Jennifer
authors.initials: J
authors.lastname: Aguilan
authors.forename: William R
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authors.lastname: Alley
authors.forename: Friederich
authors.initials: F
authors.lastname: Altmann
authors.forename: Lauren E
authors.initials: LE
authors.lastname: Ball
authors.forename: Lipika
authors.initials: L
authors.lastname: Basumallick
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authors.lastname: Bazemore-Walker
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authors.initials: H
authors.lastname: Behnken
authors.forename: Michael A
authors.initials: MA
authors.lastname: Blank
authors.forename: Kristy J
authors.initials: KJ
authors.lastname: Brown
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authors.initials: SC
authors.lastname: Bunz
authors.forename: Christopher W
authors.initials: CW
authors.lastname: Cairo
authors.forename: John F
authors.initials: JF
authors.lastname: Cipollo
authors.forename: Rambod
authors.initials: R
authors.lastname: Daneshfar
authors.forename: Heather
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authors.lastname: Desaire
authors.forename: Richard R
authors.initials: RR
authors.lastname: Drake
authors.forename: Eden P
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authors.lastname: Go
authors.forename: Radoslav
authors.initials: R
authors.lastname: Goldman
authors.forename: Clemens
authors.initials: C
authors.lastname: Gruber
authors.forename: Adnan
authors.initials: A
authors.lastname: Halim
authors.forename: Yetrib
authors.initials: Y
authors.lastname: Hathout
authors.forename: Paul J
authors.initials: PJ
authors.lastname: Hensbergen
authors.forename: David M
authors.initials: DM
authors.lastname: Horn
authors.forename: Deanna
authors.initials: D
authors.lastname: Hurum
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Jabs
authors.forename: Göran
authors.initials: G
authors.lastname: Larson
authors.forename: Mellisa
authors.initials: M
authors.lastname: Ly
authors.forename: Benjamin F
authors.initials: BF
authors.lastname: Mann
authors.forename: Kristina
authors.initials: K
authors.lastname: Marx
authors.forename: Yehia
authors.initials: Y
authors.lastname: Mechref
authors.forename: Bernd
authors.initials: B
authors.lastname: Meyer
authors.forename: Uwe
authors.initials: U
authors.lastname: Möginger
authors.forename: Christian
authors.initials: C
authors.lastname: Neusüβ
authors.forename: Jonas
authors.initials: J
authors.lastname: Nilsson
authors.forename: Milos V
authors.initials: MV
authors.lastname: Novotny
authors.forename: Julius O
authors.initials: JO
authors.lastname: Nyalwidhe
authors.forename: Nicolle H
authors.initials: NH
authors.lastname: Packer
authors.forename: Petr
authors.initials: P
authors.lastname: Pompach
authors.forename: Bela
authors.initials: B
authors.lastname: Reiz
authors.forename: Anja
authors.initials: A
authors.lastname: Resemann
authors.forename: Jeffrey S
authors.initials: JS
authors.lastname: Rohrer
authors.forename: Alexandra
authors.initials: A
authors.lastname: Ruthenbeck
authors.forename: Miloslav
authors.initials: M
authors.lastname: Sanda
authors.forename: Jan Mirco
authors.initials: JM
authors.lastname: Schulz
authors.forename: Ulrike
authors.initials: U
authors.lastname: Schweiger-Hufnagel
authors.forename: Carina
authors.initials: C
authors.lastname: Sihlbom
authors.forename: Ehwang
authors.initials: E
authors.lastname: Song
authors.forename: Gregory O
authors.initials: GO
authors.lastname: Staples
authors.forename: Detlev
authors.initials: D
authors.lastname: Suckau
authors.forename: Haixu
authors.initials: H
authors.lastname: Tang
authors.forename: Morten
authors.initials: M
authors.lastname: Thaysen-Andersen
authors.forename: Rosa I
authors.initials: RI
authors.lastname: Viner
authors.forename: Yanming
authors.initials: Y
authors.lastname: An
authors.forename: Leena
authors.initials: L
authors.lastname: Valmu
authors.forename: Yoshinao
authors.initials: Y
authors.lastname: Wada
authors.forename: Megan
authors.initials: M
authors.lastname: Watson
authors.forename: Markus
authors.initials: M
authors.lastname: Windwarder
authors.forename: Randy
authors.initials: R
authors.lastname: Whittal
authors.forename: Manfred
authors.initials: M
authors.lastname: Wuhrer
authors.forename: Yiying
authors.initials: Y
authors.lastname: Zhu
authors.forename: Chunxia
authors.initials: C
authors.lastname: Zou
mesh_terms: D002853:Chromatography, Liquid; D006023:Glycoproteins / Q000378:metabolism*; D006031:Glycosylation; D006801:Humans; D007610:Kallikreins / Q000378:metabolism*; D007753:Laboratories; D013058:Mass Spectrometry / Q000379:methods; D011134:Polysaccharides / Q000378:metabolism*; D017430:Prostate-Specific Antigen / Q000378:metabolism*; D040901:Proteomics / Q000379:methods; D015203:Reproducibility of Results
chemical_list: D006023:Glycoproteins; D011134:Polysaccharides; C549870:KLK3 protein, human; D007610:Kallikreins; D017430:Prostate-Specific Antigen