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The most sensitive AERS molecules produced Raman signals of more than 4 orders of magnitude compared to 5-ethynyl-2'-deoxyuridine (EdU). In addition, a frequency tunability of 10 distinct Raman bands was achieved by selecting different types of vibrational modes. This methodology of AERS allows for designing small-molecule Raman probes to visualize various entities in complex systems by multicolor spontaneous Raman imaging. It will open new prospects to explore innovative applications of AERS in interdisciplinary research fields.There i