# Imaging intracellular zinc by stimulated Raman scattering microscopy with a small molecule vibrational probe

## TL;DR

Scientists developed a new tool to image zinc ions in live cells using a special molecule and Raman microscopy, which could help study metal roles in biology and disease.

## Contribution

A novel small molecule vibrational probe (CSZin) enables ratiometric SRS imaging of Zn2+ and paramagnetic ions in live cells.

## Key findings

- CSZin detects Zn2+ in vitro and in live cells via SRS microscopy.
- The probe reveals changes in labile Zn2+ pools in cancer-related cell lines.
- CSZin can detect paramagnetic ions that usually quench fluorescence.

## Abstract

Advanced Raman-based techniques, particularly stimulated Raman scattering (SRS) microscopy, have emerged as valuable bioimaging tools. But advances in the development of the chemical toolbox required for detection of biologically relevant species, including metals, has lagged the rapid advances in instrumentation. To date, few responsive probes for dynamic detection of transient and low abundance species have been developed. We describe herein the design and application of CSZin, a small molecule vibrational probe for detection of Zn2+ in biological contexts. The probe, a spiropyran design with a strategically positioned nitrile, undergoes a shift in the stretching frequency of the nitrile group and an increase in the SRS intensity in response to metal-induced opening of the spiropyran. We demonstrate detection of Zn2+ ions in vitro, and the first examples of recognition-based SRS imaging of this metal in live cells. Changes in SRS intensity and ratio reveal changes in basal labile Zn2+ pools in normal and tumorigenic RWPE-1 and -2 cells that result from changes in transporter expression and cation uptake related to cancer progression. Proof-of-concept experiments demonstrate that CSZin can also respond to high concentrations of biologically relevant paramagnetic ions, enabling turn-on detection of metals that typically quench fluorescence. The modular nature of the spiropyran-based sensor may enable facile tuning of the selectivity for sensing of other target ions. This work thus paves the way for the development of a new chemical toolbox and detection strategies, complementary to fluorescence, for biological species that are challenging to image.

A spiropyran molecular probe with a strategically positioned nitrile undergoes reversible changes in its vibrational spectrum that enable ratiometric imaging of Zn2+, and paramagnetic ions, in live cells by Stimulated Raman Scattering microscopy.