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Updated: Oct 7, 2025

Combining Multiplex Fluorescence In Situ Hybridization with Fluorescent Immunohistochemistry on Fresh Frozen or Fixed Mouse Brain Sections
Published on: June 25, 2021
Hybridization chain reaction enables a unified approach to multiplexed, quantitative, high-resolution
Maayan Schwarzkopf1, Mike C Liu2, Samuel J Schulte1
1Division of Biology and Biological Engineering, California Institute of Technology, Pasadena, CA 91125, USA.
Abstract:
RNA in situ hybridization based on the mechanism of the hybridization chain reaction (HCR) enables multiplexed, quantitative, high-resolution RNA imaging in highly autofluorescent samples, including whole-mount vertebrate embryos, thick brain slices and formalin-fixed paraffin-embedded tissue sections. Here, we extend the benefits of one-step, multiplexed, quantitative, isothermal, enzyme-free HCR signal amplification to immunohistochemistry, enabling accurate and precise protein relative quantitation with subcellular resolution in an anatomical context. Moreover, we provide a unified framework for simultaneous quantitative protein and RNA imaging with one-step HCR signal amplification performed for all target proteins and RNAs simultaneously.
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