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A hybridization-chain-reaction-based method for amplifying immunosignals

Nature Methods volume 15, pages 275278 (2018) | Download Citation

Abstract

Immunosignal hybridization chain reaction (isHCR) combines antibody–antigen interactions with hybridization chain reaction (HCR) technology, which results in amplification of immunofluorescence signals by up to two to three orders of magnitude with low background. isHCR's highly modular and easily adaptable design enables the technique to be applied broadly, and we further optimized its use in multiplexed imaging and in state-of-the-art tissue expansion and clearing techniques.

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Acknowledgements

We thank R. Vale (UCSF, San Francisco, California, USA) for the pHR-scFv-GCN4-sfGFP-GB1-NLS-dWPRE plasmid, K. Deisseroth (Stanford University, Stanford, California, USA) for the pAAV-EF1α-DIO-hChR2(H134R)-mCherry plasmid, and J.H. Snyder for manuscript editing. M.L. is supported by China MOST (grants 2012YQ03026005, 2013ZX0950910, and 2015BAI08B02), NNSFC (grants 91432114 and 91632302), and the Beijing Municipal Government.

Author information

Affiliations

  1. School of Life Sciences, Peking University, Beijing, China.

    • Rui Lin
  2. National Institute of Biological Sciences (NIBS), Beijing, China.

    • Rui Lin
    • , Qiru Feng
    • , Peng Li
    • , Ping Zhou
    • , Ruiyu Wang
    • , Zhe Liu
    • , Zhiqiang Wang
    • , Xiangbing Qi
    • , Nan Tang
    • , Feng Shao
    •  & Minmin Luo
  3. Peking University-Tsinghua University-NIBS Joint Graduate Program, NIBS, Beijing, China.

    • Rui Lin
  4. School of Life Sciences, Tsinghua University, Beijing, China.

    • Qiru Feng
    •  & Minmin Luo

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Contributions

R.L. and M.L. conceived of the application of HCR to immunosignal amplification. R.L. and P.Z. designed and constructed the clones. F.S., P.Z., and P.L. designed the S. Typhimurium infection experiments, and R.L. and P.L. performed S. Typhimurium infection experiments. R.L. and P.L. performed protein purification and western blotting. X.Q. and Z.W. designed and synthesized the chemical linkers. Q.F. packaged the AAV vectors. Z.L., R.L., and N.T. designed the tissue-clearing experiments, and R.L., Q.F., and Z.L. performed the tissue-clearing experiments. R.L., R.W., and M.L. analyzed the data. R.L. and M.L. wrote the manuscript.

Competing interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to Minmin Luo.

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DOI

https://doi.org/10.1038/nmeth.4611

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