Peptide and protein quantification using automated immuno-MALDI (iMALDI)

Huiyan Li, Robert Popp, Bjorn Frohlich, Michael X. Chen, Christoph H. Borchers

Research output: Contribution to journalArticlepeer-review

7 Citations (Scopus)

Abstract

Mass spectrometry (MS) is one of the most commonly used technologies for quantifying proteins in complex samples, with excellent assay specificity as a result of the direct detection of the mass-to-charge ratio of each target molecule. However, MS-based proteomics, like most other analytical techniques, has a bias towards measuring high-abundance analytes, so it is challenging to achieve detection limits of low ng/mL or pg/ mL in complex samples, and this is the concentration range for many disease-relevant proteins in biofluids such as human plasma. To assist in the detection of low-abundance analytes, immuno-enrichment has been integrated into the assay to concentrate and purify the analyte before MS measurement, significantly improving assay sensitivity. In this work, the immuno-Matrix-Assisted Laser Desorption/Ionization (iMALDI) technology is presented for the quantification of proteins and peptides in biofluids, based on immuno-enrichment on beads, followed by MALDI-MS measurement without prior elution. The anti-peptide antibodies are functionalized on magnetic beads, and incubated with samples. After washing, the beads are directly transferred onto a MALDI target plate, and the signals are measured by a MALDI-Time of Flight (MALDI-TOF) instrument after the matrix solution has been applied to the beads. The sample preparation procedure is simplified compared to other immuno-MS assays, and the MALDI measurement is fast. The whole sample preparation is automated with a liquid handling system, with improved assay reproducibility and higher throughput. In this article, the iMALDI assay is used for determining the peptide angiotensin I (Ang I) concentration in plasma, which is used clinically as readout of plasma renin activity for the screening of primary aldosteronism (PA).

Original languageEnglish
Article numbere55933
JournalJournal of Visualized Experiments
Volume2017
Issue number126
DOIs
Publication statusPublished - Aug 2017
Externally publishedYes

Keywords

  • Automation
  • Chemistry
  • IMALDI
  • Immuno-enrichment
  • Immuno-MALDI
  • Issue 126
  • Mass spectrometry
  • Peptides
  • Protein quantification

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