Hybridization chain reaction enables a unified approach to multiplexed, quantitative, high-resolution immunohistochemistry and in situ hybridization

ABSTRACT RNA in situ hybridization (RNA-ISH) based on the mechanism of 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 (FFPE) tissue sections. Here, we extend the benefits of 1-step, multiplexed, quantitative, isothermal, enzyme-free HCR signal amplification to immunohistochemistry (IHC), enabling accurate and precise protein relative quantitation with subcellular resolution in an anatomical context. More-over, we provide a unified framework for simultaneous quantitative protein and RNA imaging with 1-step HCR signal amplification performed for all target proteins and RNAs simultaneously. SUMMARY Signal amplification based on the mechanism of hybridization chain reaction enables multiplexed, quantitative, high-resolution imaging of protein and RNA targets in highly autofluorescent tissues.