ArticleScientific reports2017
Highly sensitive and specific protein detection via combined capillary isoelectric focusing and proximity ligation.
Article in Scientific reports, 2017. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 7 papers.
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Who cites it
7 citing papers in PubMed.
- Proximity Ligation Assay as a Tool for Antibody Validation in Human Tissues.The journal of histochemistry and cytochemistry : official journal of the Histochemistry Society · 2020Article
- Article
- Differentiation of Essential Oils Using Nanofluidic Protein Post-Translational Modification Profiling.Molecules (Basel, Switzerland) · 2019Article
- Chaperome heterogeneity and its implications for cancer study and treatment.The Journal of biological chemistry · 2019Review
- Plasma Protein Profiling Reveal Osteoprotegerin as a Marker of Prognostic Impact for Colorectal Cancer.Translational oncology · 2018Article
- Detection of the Cell Cycle-Regulated Negative Feedback Phosphorylation of Mitogen-Activated Protein Kinases in Breast Carcinoma using Nanofluidic Proteomics.Scientific reports · 2018Article
- Improved efficiency of in situ protein analysis by proximity ligation using UnFold probes.Scientific reports · 2018Article
Corrections and comments
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Authors and funding
10 authors.
Funding
No grant is acknowledged in the PubMed record.
Abstract
Detection and quantification of proteins and their post-translational modifications are crucial to decipher functions of complex protein networks in cell biology and medicine. Capillary isoelectric focusing together with antibody-based detection can resolve and identify proteins and their isoforms with modest sample input. However, insufficient sensitivity prevents detection of proteins present at low concentrations and antibody cross-reactivity results in unspecific detection that cannot be distinguished from bona fide protein isoforms. By using DNA-conjugated antibodies enhanced signals can be obtained via rolling circle amplification (RCA). Both sensitivity and specificity can be greatly improved in assays dependent on target recognition by pairs of antibodies using in situ proximity ligation assays (PLA). Here we applied these DNA-assisted RCA techniques in capillary isoelectric focusing to resolve endogenous signaling transducers and isoforms along vascular endothelial growth factor (VEGF) signaling pathways at concentrations too low to be detected in standard assays. We also demonstrate background rejection and enhanced specificity when protein detection depended on binding by pairs of antibodies using in situ PLA, compared to assays where each antibody preparation was used on its own.
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