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Analysis of Histone Antibody Specificity with Peptide Microarrays
Published on: August 1, 2017
Peptide microarrays for profiling of modification state-specific antibodies
Johannes Zerweck1, Antonia Masch, Mike Schutkowski
1JPT Peptide Technologies GmbH, Volmerstr. 5, D-12489 Berlin, Germany.
Methods in Molecular Biology (Clifton, N.J.)
|April 21, 2009
Summary
Generic antibodies for detecting protein phosphorylation and acetylation show unexpected specificity. Researchers found these tools have distinct subsite preferences and cross-reactivity, impacting signal transduction and proteomics research.
Area of Science:
- Biochemistry
- Molecular Biology
- Proteomics
Background:
- Post-translational modifications like phosphorylation regulate crucial cellular processes.
- Accurate detection of modified amino acid residues is vital for signal transduction research.
- Pan-specific antibodies are widely used but their precise reactivity is often assumed.
Purpose of the Study:
- To comprehensively characterize commercially available generic anti-phosphopeptide and anti-acetyllysine antibodies.
- To assess the specificity and potential cross-reactivity of these widely used antibodies.
Main Methods:
- Utilized high-content phosphopeptide microarrays.
- Employed microarrays displaying acetyllysine-containing peptides.
- Performed comprehensive characterization of antibody binding.
Main Results:
- Demonstrated distinct subsite specificity for anti-phosphopeptide antibodies.
- Revealed cross-reactivity patterns among tested antibodies.
- Characterized the reactivity of anti-acetyllysine antibodies on specific peptide arrays.
Conclusions:
- Generic antibodies exhibit more specific binding than previously assumed.
- Understanding antibody subsite specificity is crucial for accurate interpretation of proteomics and signal transduction data.
- Re-evaluation of antibody validation methods is necessary for reliable research outcomes.
