Non-enzymatic, high-gain signal amplification methods with single-cell, single-molecule resolution are in great need. We present click-amplifying FISH (clampFISH) for the fluorescent detection of RNA that combines the specificity of oligonucleotides with bioorthogonal click chemistry in order to achieve high specificity and extremely high-gain (>400x) signal amplification. We show that clampFISH signal enables detection with low magnification microscopy and separation of cells by RNA levels via flow cytometry. Additionally, we show that the modular design of clampFISH probes enables multiplexing, that the locking mechanism prevents probe detachment in expansion microscopy, and that clampFISH works in tissue samples.
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