Dual-phase stimulated Raman scattering microscopy for real-time two-color imaging

Ruoyu He,Minbiao Ji,Shenghong Ma,Lili Zhang,Dan Ye,Yongkui Xu,Xu Wang

doi:10.1364/optica.4.000044

Abstract

Label-free histology with chemical contrast has great potential for rapid intraoperative diagnosis. Two-color stimulated Raman scattering (SRS) microscopy has shown success in label-free digital histology with diagnostic results similar to those of hematoxylin and eosin stain. However, achieving real-time two-color SRS imaging has been a challenge. We have precisely engineered the pulse profiles of Stokes beams, and fully utilized in-phase (X) and quadrature (Y) outputs of a phase-sensitive lock-in amplifier to realize simultaneous two-color imaging. Such a method could reach the maximum speed as in single-color SRS, and has proven its robustness and advantages in real-time histology as well as in vivo imaging of live animals, both in transmission and epi modes. Moreover, this method could be conveniently adapted to other pump–probe-based microscopes.

Highlights

As one of the coherent Raman scattering (CRS) imaging techniques, stimulated Raman scattering (SRS) microscopy has demonstrated unique capabilities in quantitative chemical analysis, live cell drug delivery, DNA imaging, and tumor detection [1,2,3,4,5,6,7,8,9]
Real-time histological imaging with multichemical contrast has never been achieved in live animals in epi mode, which is a key step for the clinical translation of SRS microscopy
We have previously demonstrated the potential of two-color SRS microscopy as a label-free histological tool to delineate tumor from normal tissues [5,32]

Summary

Introduction

As one of the coherent Raman scattering (CRS) imaging techniques, stimulated Raman scattering (SRS) microscopy has demonstrated unique capabilities in quantitative chemical analysis, live cell drug delivery, DNA imaging, and tumor detection [1,2,3,4,5,6,7,8,9]. Dual-phase stimulated Raman scattering microscopy for real-time two-color imaging We present a compact design of parallel two-color SRS microscopy with the two orthogonal outputs of a dual-phase lock-in amplifier (LIA).

Results

Conclusion