A ratiometric Raman probe for live-cell imaging of hydrogen sulfide in mitochondria by stimulated Raman scattering

Abstract

Stimulated Raman Scattering (SRS) coupled with alkyne tags has been an emerging imaging technique to visualize small-molecule species with high sensitivity and specificity. Here we describe the development of a ratiometric Raman probe for visualizing hydrogen sulfide (H2S) species in living cells as the first alkyne-based sensor for SRS microscopy. This probe uses an azide unit as a selective reactive site, and it targets mitochondria with high specificity. The SRS ratiometric images show a strong response to H2S level changes in living cells.

A new class of ratiometric small molecule intracellular pH sensors for Raman microscopy

“…In addition, diffuse absorption and emission spectra hinder the analysis of multiple fluorescent probes simultaneously (multiplexing), since spectral overlap limits the number of resolvable features within the spectra (especially where ratiometric sensors are used). 20,21 Intracellular pH sensing has also been achieved by Surface Enhanced Raman Scattering (SERS) using functionalised metal nanoparticles. [22][23][24][25][26] These nanoparticles bear a surface coating of organic molecules with pH-dependent Raman spectra, which are detectable inside cells with excellent sensitivity, owing to the SERS effect.…”

“…1B). 20 This sensor contained an azide group in conjugation with a bisarylbutadiyne scaffold which was reduced to the corresponding aniline in the presence of H 2 S. This chemical transformation induced a Raman shift change in the alkyne band, enabling the detection of intracellular H 2 S by SRS microscopy. Whilst this clearly demonstrates that Raman microscopy can be used to develop intracellular sensors, it relies on an irreversible chemical process rendering the detection semi-quantitative.…”

Recent progress of surface-enhanced Raman spectroscopy for subcellular compartment analysis

“…Mitochondrion has a membrane potential of nearly -180 mV, and such a negative potential allows positively charged nanomaterials in the mitochondria autonomously 56 . Accordingly, many mitochondrion-targeting synthetic materials were designed by enriching them with positive charges, e.g., a positively ratiometric Raman probe was prepared to achieve the selective mitochondria targeting and live-cell imaging of hydrogen sulfide in cancer cells 57 . Although passive targeting shows apparent organelle-targeting to a certain degree, it is still limited in its applicability for all kinds of NPs.…”

Imaging the invisible—Bioorthogonal Raman probes for imaging of cells and tissues

“…They detected the so-called MitoBADY in living HeLa cells after only 5 minutes indicating a rapid uptake, while the signal was coinciding with cytochrome c. This opens the door to study mitochondrial dynamics and endogeneous cellular mechanisms simultaneously recorded within the same Raman spectrum. Zeng et al designed a ratiometric Raman probe for mitochondria, which is sensing hydrogen sulfide (H 2 S) changes in living cells [94]. The probe has a reactive site, which in the presence of H 2 S experiences a reduction from an azide to an amine group causing a Raman frequency shift by 9 cm −1 .…”

“…A significant increase in the ratio can be seen in the presence of NaSH. Reproduced from reference Zeng et al [94] with permission from The Royal Society of Chemistry. C, Spontaneous Raman spectra of living MCF-7 cells incubated with the indicated labeled substrates (AA, amino acid; PA, palmitic acid; Glc, glucose) are shown.…”