Third-generation in situ hybridization chain reaction: multiplexed, quantitative, sensitive, versatile, robust
By Harry M. T. Choi, Maayan Schwarzkopf, M. Fornace, Aneesha Acharya, Georgios Artavanis, J. Stegmaier, Alexandre Cunha, Niles A. Pierce. Development, 2018
1,269 citations recorded by the source corpus.
ABSTRACT In situ hybridization based on the mechanism of the hybridization chain reaction (HCR) has addressed multi-decade challenges that impeded imaging of mRNA expression in diverse organisms, offering a unique combination of multiplexing, quantitation, sensitivity, resolution and versatility. Here, with third-generation in situ HCR, we augment these capabilities using probes and amplifiers that combine to provide automatic background suppression throughout the protocol, ensuring that reagents will not generate amplified background even if they bind non-specifically within the sample. Automatic background suppression dramatically enhances performance and robustness, combining the benefits of a higher signal-to-background ratio with the convenience of using unoptimized probe sets for new targets and organisms. In situ HCR v3.0 enables three multiplexed quantitative analysis modes: (1) qHCR imaging – analog mRNA relative quantitation with subcellular resolution in the anatomical context of whole-mount vertebrate embryos; (2) qHCR flow cytometry – analog mRNA relative quantitation for high-throughput expression profiling of mammalian and bacterial cells; and (3) dHCR imaging – digital mRNA absolute quantitation via single-molecule imaging in thick autofluorescent samples. Highlighted Article: In situ hybridization chain reaction (HCR) v3.0 exploits automatic background suppression to enable multiplexed quantitative mRNA imaging and flow cytometry with dramatically enhanced performance and ease of use.
Subfields: Molecular Biology
View Paper: https://www.semanticscholar.org/paper/905932a64d04b1def045784f9501085b43416d04
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In Molecular Biology
Citations in the corpus
- Non-Canonical Odor Coding in the Mosquito (2020, methodology): “RNA was detected in whole mount antenna and maxillary palp using the hybridization chain reaction (HCR) technique as previously described (Choi et al., 2018) with modifications.”