Switchable stimulated Raman scattering microscopy with photochromic vibrational probes

Jianpeng Ao,Xiaofeng Fang,Hyunchul Kang,Jiwei Ling,Sungnam Park,Xianchong Miao,Changfeng Wu,Minbiao Ji

doi:10.1038/s41467-021-23407-2

Abstract

Photochromic probes with reversible fluorescence have revolutionized the fields of single molecule spectroscopy and super-resolution microscopy, but lack sufficient chemical specificity. In contrast, Raman probes with stimulated Raman scattering (SRS) microscopy provides superb chemical resolution for super-multiplexed imaging, but are relatively inert. Here we report vibrational photochromism by engineering alkyne tagged diarylethene to realize photo-switchable SRS imaging. The narrow Raman peak of the alkyne group shifts reversibly upon photoisomerization of the conjugated diarylethene when irradiated by ultraviolet (UV) or visible light, yielding “on” or “off” SRS images taken at the photoactive Raman frequency. We demonstrated photo-rewritable patterning and encryption on thin films, painting/erasing of cells with labelled alkyne-diarylethene, as well as pulse-chase experiments of mitochondria diffusion in living cells. The design principle provides potentials for super-resolution microscopy, optical memories and switches with vibrational specificity.

Highlights

Photochromic probes with reversible fluorescence have revolutionized the fields of single molecule spectroscopy and super-resolution microscopy, but lack sufficient chemical specificity
While the electronic transitions of fluorescent molecules have the advantage of ultrabrightness up to single molecule sensitivity, the intrinsic broad/ overlapping spectral feature has limited the resolving power of probe species
stimulated Raman scattering (SRS) microscopy was initially developed as a label-free chemical imaging technique without the need of exogenous labeling a DTE-Ph-Mito open

Summary

Introduction

Photochromic probes with reversible fluorescence have revolutionized the fields of single molecule spectroscopy and super-resolution microscopy, but lack sufficient chemical specificity. The narrow Raman peak of the alkyne group shifts reversibly upon photoisomerization of the conjugated diarylethene when irradiated by ultraviolet (UV) or visible light, yielding “on” or “off” SRS images taken at the photoactive Raman frequency. Further coupled with engineered Raman probes, SRS has demonstrated various imaging capabilities competing with fluorescence based microscopies[14,15] These include super-multiplexed imaging using polyynes and alkyne-tagged dyes to resolve more than 20 labels simultaneously[16,17], and single molecule vibrational sensitivity with stimulated Raman excited fluorescence[18,19]. We characterized the basic vibrational photo-switching properties of the engineered Raman probes, and applied them to rewritable information patterning, as well as controlled activation, erasing and chasing in living cells with SRS microscopy

Methods

Results

Conclusion