Raman scattering
provides stable narrow-banded signals that potentially
allow for multicolor microscopic imaging. The major obstacle for the
applications of Raman spectroscopy and microscopy is the small cross
section of Raman scattering that results in low sensitivity. Here,
we report a new concept of azo-enhanced Raman scattering (AERS) by
designing the intrinsic molecular structures using resonance Raman
and concomitant fluorescence quenching strategies. Based on the selection
of vibrational modes and the enhancing unit of azobenzenes, we obtained
a library of AERS molecules with specific Raman signals in the fingerprint
and silent frequency regions. The spectral characterization and molecular
simulation revealed that the azobenzene unit conjugated to the vibrational
modes significantly enhanced Raman signals due to the mechanism of
extending the conjugation system, coupling the electronic–vibrational
transitions, and improving the symmetry of vibrational modes. The
nonradiative decay of azobenzene from the excited state quenched the
commitment fluorescence, thus providing a clean background for identifying
Raman scattering. 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.