HCR hairpin
HCR Citation Notes

For citation, please select from the list below as appropriate for your application:

  • 10-plex HCR spectral imaging
    HCR spectral imaging of any combination of 10 RNA, protein, or protein:protein targets with 1-step quantitative HCR signal amplification for all targets simultaneously (Schulte et al., 2024a).
  • HCR protein:protein complex imaging
    Multiplexed, quantitative, high-resolution imaging of protein:protein complexes with/without HCR IF and/or HCR RNA-FISH with simultaneous HCR signal amplification for all protein:protein, protein, and RNA targets (Schulte et al., 2024b)
    A unified approach to multiplexed, quantitative, high-resolution RNA fluorescence in situ hybridization (RNA-FISH) and protein immunofluorescence (IF) with 1-step quantitative enzyme-free HCR signal amplification performed for all RNA and protein targets simultaneously (Schwarzkopf et al., 2021)
  • HCR IF
    Multiplexed, quantitative, high-resolution protein immunofluorecence (IF) in highly autofluorescent samples (e.g., FFPE brain tissue sections) (Schwarzkopf et al., 2021)
  • HCR RNA-FISH (v3.0)
    Automatic background suppression for dramatically enhanced performance (signal-to-background and quanti- tative precision) and ease-of-use (no probe set optimization for new targets) (Choi et al., 2018). v3.0 supports three quantitative modes:
    qHCR imaging: analog mRNA relative quantitation with subcellular resolution.
    qHCR flow cytometry: analog mRNA relative quantitation for high-throughput single-cell analysis.
    dHCR imaging: digital mRNA absolute quantitation.
    Protocols for v3.0 in diverse organisms are adapted from the zoo paper.
    Software: Dot Analysis 1.0 package.
  • Zoo paper
    Protocols for multiplexed mRNA imaging in diverse sample types (Choi et al., 2016):
    bacteria in suspension
    FFPE human tissue sections
    generic sample in suspension
    generic sample on slide
    whole-mount chicken embryos
    whole-mount fruit fly embryos
    whole-mount mouse embryos
    whole-mount sea urchin embryos
    whole-mount worm larvae
    whole-mount zebrafish embryos and larvae
  • Single-molecule HCR imaging
    Single-molecule mRNA imaging in thick autofluorescent samples (e.g., 0.5 mm adult mouse brain sections) (Shah et al., 2016).

  • Multiplexed quantitative HCR (qHCR) northern blots
    Simultaneous quantification of RNA target size and abundance with signal amplification for up to 5 target RNAs (Schwarzkopf & Pierce, 2016).
  • In situ HCR RNA-FISH (v2.0)
    2nd generation in situ HCR technology (v2.0) using DNA probes and DNA HCR amplifiers: 10× increase in signal, 10× reduction in cost, dramatic increase in reagent durability (Choi et al., 2014).
  • Shielded covalent (SC) probes
    Highly selective covalent capture of DNA and RNA targets, overcoming the longstanding tradeoff between selectivity and durable target capture (Vieregg et al., 2013).
  • HCR RNA-FISH (v1.0)
    1st generation in situ HCR technology (v1.0) using RNA probes and RNA HCR amplifiers: multiplexed mRNA imaging in whole-mount vertebrate embryos with simultaneous signal amplification for up to 5 target mRNAs (Choi et al., 2010).
  • Hybridization chain reaction (HCR) mechanism
    The hybridization chain reaction (HCR) mechanism enables isothermal enzyme-free molecular signal amplification (Dirks & Pierce, 2004).

HCR Technology References
  • Choi, H.M.T., Beck, V.A., & Pierce, N.A. (2014). Next-generation in situ hybridization chain reaction: higher gain, lower cost, greater durability. ACS Nano, 8(5):4284-4294. (pdf, supp info, movie)
  • Choi, H.M.T., Calvert, C.R., Husain, N., Huss, D., Barsi, J.C., Deverman, B.E., Hunter, R.C., Kato, M., Lee, S.M., Abelin, A.C.T., Rosenthal, A.Z., Akbari, O.S., Li, Y., Hay, B.A., Sternberg, P.W., Patterson, P.H., Davidson, E.H., Mazmanian, S.K., Prober, D.A., van de Rijn, M., Leadbetter, J.R., Newman, D.K., Readhead, C., Bronner, M.E., Wold, B., Lansford, R., Sauka-Spengler, T., Fraser, S.E., & Pierce, N.A. (2016). Mapping a multiplexed zoo of mRNA expression. Development, 143:3632-3637. (pdf, supp info, fruit fly movie, sea urchin movie, zebrafish movie, chicken movie, mouse movie)
  • Choi, H.M.T., Schwarzkopf, M., Fornace, M.E., Acharya, A., Artavanis, G., Stegmaier, J., Cunha, A., & Pierce, N.A. (2018). Third-generation in situ hybridization chain reaction: multiplexed, quantitative, sensitive, versatile, robust. Development, 145, dev165753. (pdf, supp info, Dot Analysis 1.0 package)
  • Dirks, R.M., & Pierce, N.A. (2004). Triggered amplification by hybridization chain reaction. Proc Natl Acad Sci USA, 101(43), 15275–15278. (pdf)
  • Schulte, S.J., Shin, B., Rothenberg, E.V., & Pierce, N.A. (2024b). Multiplex, quantitative, high-resolution imaging of protein:protein complexes via hybridization chain reaction. ACS Chem Biol, 19(2), 280–288. (pdf, supp info)
  • Schwarzkopf, M., Liu, M. C., Schulte, S. J., Ives, R., Husain, N., Choi, H. M. T., & Pierce, N. A. (2021). Hybridization chain reaction enables a unified approach to multiplexed, quantitative, high-resolution immunohistochemistry and in situ hybridization. Development, 148(22), dev199847. (pdf, supp info)
  • Schwarzkopf, M., & Pierce, N.A. (2016). Multiplexed miRNA northern blots via hybridization chain reaction. Nucleic Acids Res, 44(15), e129.
    (pdf, supp info)
  • Shah, S., Lubeck, E., Schwarzkopf, M., He, T.-F., Greenbaum, A., Sohn, C.H., Lignell, A., Choi, H.M.T., Gradinaru, V., Pierce, N.A., & Cai, L. (2016). Single-molecule RNA detection at depth via hybridization chain reaction and tissue hydrogel embedding and clearing. Development, 143:2862-2867. (pdf, supp info, movie 1, movie 2, movie 3)
  • Vieregg, J.R., Nelson, H.M., Stoltz, B.M., & Pierce, N.A. (2013). Selective nucleic acid capture with shielded covalent probes. J Am Chem Soc, 135(26), 9691–9699.
    (pdf, supp info)