Electronic Preresonance Stimulated Raman Scattering Microscopy

Abstract

Optical microscopy has generated great impact for modern research. While fluorescence microscopy provides the ultimate sensitivity, it generally lacks chemical information. Complementarily, vibrational imaging methods provide rich chemical-bond-specific contrasts. Nonetheless, they usually suffer from unsatisfying sensitivity or compromised biocompatibility. Recently, electronic preresonance stimulated Raman scattering (EPR-SRS) microscopy was reported, achieving simultaneous high detection sensitivity and superb vibrational specificity of chromophores. With newly synthesized Raman-active dyes, this method readily breaks the optical color barrier of fluorescence microscopy and is well-suited for supermultiplex imaging in biological samples. In this Perspective, we first review previous utilizations of electronic resonance in various Raman spectroscopy and microscopy. We then discuss the physical origin and uniqueness of the electronic preresonance region, followed by quantitative analysis of the enhancement factors involved in EPR-SRS microscopy. On this basis, we provide an outlook for future development as well as the broad applications in biophotonics.

Polydiacetylene-based ultrastrong bioorthogonal Raman probes for targeted live-cell Raman imaging

“…Since PDDA had a maximum absorption at 476 nm in DMSO (Supplementary Fig. 6b), the Raman excitation at 488 nm, a wavelength close to its absorption maximum, was able to further enhance the Raman intensity through pre-resonance Raman scattering 48,49 , resulting in a tremendously high RIE value of 2.3 × 10 4 (C≡C bond-normalized).…”

Plasmon-enhanced stimulated Raman scattering microscopy with single-molecule detection sensitivity

“…Yet, the imaging sensitivity of SRS microscopy is limited to ~10 mM for chemical bonds such as the C-H vibrations in cell membranes 22,28 . Min and coworkers recently reported electronic pre-resonance SRS achieving sub-micromolar-sensitivity detection for chromophores having a Raman cross-section 10 3 or 10 4 times larger than endogenous biomolecules 29,30 .…”

Electronic Resonant Stimulated Raman Scattering Micro-Spectroscopy

“…This value is at least 100 times stronger than the effective cross sections determined in epr-SRS! 15 . To retrieve the vibrational feature in er-SRS, it is necessary to suppress the large background from chromophores first.…”

“…In our study of combining electronic resonance and SRS, when we reached the rigorously resonant scenario (| ω 0 − ω pump | < Γ, Γ ≈ 700 cm −1 is the homogeneous linewidth), only overwhelming electronic background without any vibrational features was observed for the infrared dye IR895 14 . This led us take a step back to the pre-resonance region (2Γ < ω 0 − ω pump < 6Γ) which is a sweet spot without background influence but still retain sufficiently high signal enhancement factor (EF) up to 2 × 10 5 for Atto740 through electronic pre-resonance effect 14–15 . However, EF as large as 6–7 orders of magnitude have been reported in literature for rigorous electronic resonance region 4–5, 9–11, 16 .…”