In situ protein detection with enhanced specificity using DNA-conjugated antibodies and proximity ligation.
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Zieba A, Ponten F, Uhlén M, Landegren U
Mod. Pathol. 31 (2) 253-263 [2018-02-00; online 2017-09-22]
Antibodies are important tools in anatomical pathology and research, but the quality of in situ protein detection by immunohistochemistry greatly depends on the choice of antibodies and the abundance of the targeted proteins. Many antibodies used in scientific research do not meet requirements for specificity and sensitivity. Accordingly, methods that improve antibody performance and produce quantitative data can greatly advance both scientific investigations and clinical diagnostics based on protein expression and in situ localization. We demonstrate here protocols for antibody labeling that allow specific protein detection in tissues via bright-field in situ proximity ligation assays, where each protein molecule must be recognized by two antibodies. We further demonstrate that single polyclonal antibodies or purified serum preparations can be used for these dual recognition assays. The requirement for protein recognition by pairs of antibody conjugates can significantly improve specificity of protein detection over single-binder assays.
Affinity Proteomics Uppsala [Technology development]
NGI Stockholm (Genomics Applications) [Service]
NGI Stockholm (Genomics Production) [Service]
National Genomics Infrastructure [Service]
PLA and Single Cell Proteomics [Technology development]
Tissue Profiling [Collaborative]
PubMed 28937142
DOI 10.1038/modpathol.2017.102
Crossref 10.1038/modpathol.2017.102
pii: modpathol2017102