ArticleAnalytical chemistry2019
Restricted Proteolysis and LC-MS/MS To Evaluate the Orientation of Surface-Immobilized Antibodies.
Article in Analytical chemistry, 2019. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 4 papers.
What it found
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The abstract states no effect estimate the extractor could read, or names no intervention and outcome on the map, so this paper lights no cell and moves no belief. It is still indexed, cited and linked below.
The trial behind it
Trials whose registry record cites this paper, or whose number appears in the abstract. A trial that started after this paper was published is citing it as background, not reporting it.
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Who cites it
4 citing papers in PubMed.
- A robust and scalable immunoadsorbent platform based on the high-affinity M03-IgG antibody for targeted IL-6 Removal in cytokine storm.Frontiers in immunology · 2026Article
- A Concise Review of the Control and Assessment of Magnetic Affinity Particle Assembly for Live Cell Analyses: State of the Art and Challenges.Materials (Basel, Switzerland) · 2025Review
- Analytical methods of antibody surface coverage and orientation on bio-functionalized magnetic beads: application to immunocapture of TNF-α.Analytical and bioanalytical chemistry · 2021Article
- Organ-Specific Screening for Protein Damage Using Magnetic Bead Bioreactors and LC-MS/MS.Analytical chemistry · 2020Article
Corrections and comments
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Authors and funding
5 authors.
Funding
Abstract
The molecular orientation of antibodies immobilized on solid surfaces plays a significant role in the sensitivity of immunoassays and efficiency of protein isolation using antibody-decorated nanoparticles. Optimally, nearly all antibody binding sites should be available to bind. Here we report for the first time an LC-MS/MS approach to probe antibody orientation directly, utilizing sterically restricted proteolysis. Trypsin-decorated magnetic beads (MBs, 1.5 μm) were much larger than average antibody-free areas (55 × 55 nm) of oriented antibodies on MBs, restricting proteolysis to mainly Fab regions. Randomly attached antibodies on MB surfaces served as controls. The tryptic-hydrolyzed peptides were quantified using LC-MS/MS peptide analysis as markers for average positions of Fc and Fab of antibodies on the beads. Different patterns of digestion rates were found due to proteolysis of the oriented and nonoriented antibodies on MBs. For oriented antibodies, the peptides from outer Fab regions gave a much higher digestion rate than those from Fc regions, while for randomly immobilized antibodies digestion rates for Fab and Fc peptides were similar. This novel approach is a useful and convenient tool to characterize antibody orientation for immunoassays and other applications. The relative degree of orientation can be assessed using a metric R
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Registered trials
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