High-throughput MS-based protein phenotyping: Application to haptoglobin

Kemmons A. Tubbs, Urban A. Kiernan, Eric E. Niederkofler, Dobrin Nedelkov, Allan L. Bieber, Randall W. Nelson

Research output: Contribution to journalArticle

17 Citations (Scopus)

Abstract

A high-throughput affinity capture and reduction approach was developed for phenotype and post-translational modification analysis of a complexed globular protein, haptoglobin (Hp), directly from human plasma. Hp was selectively retrieved utilizing anti-Hp antibodies immobilized onto affinity pipette tips, eluted onto a formatted mass spectrometer target for reduction of Hp α-chains (Hpα1 and Hpα2) and subjected to subsequent MALDI-MS analysis. The affinity capture and reduction approach was originally developed from a pre-extraction reduction methodology that was optimized to an affinity capture post-reduction technique for intact Hp α-chain variant analysis, phenotype classification and ensuing post-translational variant detection. Three common Hp phenotypes (1-1, 2-1 and 2-2) were assigned according to detection of Hpα1 and/or Hpα2 reduced intact chain(s) average mass(es). The affinity capture post-reduction approach was scaled for high-throughput Hp α-chain phenotype analysis from a normal plasma cohort. The entire sample cohort was successfully analyzed and phenotyped using the developed approach. Additionally, Hp post-translational variants were detected and assigned via accurate MS analyses. The results of this study suggest use of the methodology in future analyses of other similarly complexed proteins and in normal versus disease cohort population proteomics studies.

Original languageEnglish (US)
Pages (from-to)5002-5007
Number of pages6
JournalProteomics
Volume5
Issue number18
DOIs
StatePublished - Dec 2005

Fingerprint

Haptoglobins
Throughput
Proteins
Phenotype
Post and Core Technique
Immobilized Antibodies
Plasma (human)
Matrix-Assisted Laser Desorption-Ionization Mass Spectrometry
Post Translational Protein Processing
Proteomics
Mass spectrometers
Anti-Idiotypic Antibodies
Plasmas
Population

Keywords

  • Haptoglobin
  • High-throughput
  • MS
  • Protein phenotyping

ASJC Scopus subject areas

  • Molecular Biology
  • Genetics

Cite this

Tubbs, K. A., Kiernan, U. A., Niederkofler, E. E., Nedelkov, D., Bieber, A. L., & Nelson, R. W. (2005). High-throughput MS-based protein phenotyping: Application to haptoglobin. Proteomics, 5(18), 5002-5007. https://doi.org/10.1002/pmic.200500176

High-throughput MS-based protein phenotyping : Application to haptoglobin. / Tubbs, Kemmons A.; Kiernan, Urban A.; Niederkofler, Eric E.; Nedelkov, Dobrin; Bieber, Allan L.; Nelson, Randall W.

In: Proteomics, Vol. 5, No. 18, 12.2005, p. 5002-5007.

Research output: Contribution to journalArticle

Tubbs, KA, Kiernan, UA, Niederkofler, EE, Nedelkov, D, Bieber, AL & Nelson, RW 2005, 'High-throughput MS-based protein phenotyping: Application to haptoglobin', Proteomics, vol. 5, no. 18, pp. 5002-5007. https://doi.org/10.1002/pmic.200500176
Tubbs KA, Kiernan UA, Niederkofler EE, Nedelkov D, Bieber AL, Nelson RW. High-throughput MS-based protein phenotyping: Application to haptoglobin. Proteomics. 2005 Dec;5(18):5002-5007. https://doi.org/10.1002/pmic.200500176
Tubbs, Kemmons A. ; Kiernan, Urban A. ; Niederkofler, Eric E. ; Nedelkov, Dobrin ; Bieber, Allan L. ; Nelson, Randall W. / High-throughput MS-based protein phenotyping : Application to haptoglobin. In: Proteomics. 2005 ; Vol. 5, No. 18. pp. 5002-5007.
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