Advancing the immunoaffinity platform AFFIRM to targeted measurements of proteins in serum in the pg/ml range

Research output: Contribution to journalArticle

Abstract

There is a great need for targeted protein assays with the capacity of sensitive measurements in complex samples such as plasma or serum, not the least for clinical purposes. Proteomics keeps generating hundreds of biomarker candidates that need to be transferred towards true clinical application through targeted verification studies and towards clinically applicable analysis formats. The immunoaffinity assay AFFIRM (AFFInity sRM) combines the sensitivity of recombinant single chain antibodies (scFv) for targeted protein enrichment with a specific mass spectrometry readout through selected reaction monitoring (SRM) in an automated workflow. Here we demonstrate a 100 times improved detection capacity of the assay down to pg/ml range through the use of oriented antibody immobilization to magnetic beads. This was achieved using biotin-tagged scFv coupled to streptavidin coated magnetic beads, or utilizing the FLAG tag for coupling to anti-FLAG antibody coated magnetic beads. An improved multiplexing capacity with an 11-plex setup was also demonstrated compared to a previous 3-plex setup, which is of great importance for the analysis of panels of biomarker targets.

Details

Authors
  • Anna Säll
  • Daniel Corbee
  • Sara Vikström
  • Filip Ottosson
  • Helena Persson
  • Sofia Waldemarson
Organisations
External organisations
  • KTH Royal Institute of Technology
  • Lund University
Research areas and keywords

Subject classification (UKÄ) – MANDATORY

  • Immunology in the medical area
  • Medical Laboratory and Measurements Technologies
Original languageEnglish
Article numbere0189116
JournalPLoS ONE
Volume13
Issue number2
Publication statusPublished - 2018 Feb 1
Publication categoryResearch
Peer-reviewedYes