Ludwig-Soret microscopy with vibrational photothermal effect

TL;DR

This paper introduces ViP microscopy for label-free intracellular thermophoretic imaging, revealing molecular transport behaviors driven by temperature gradients in living cells with high spatial resolution.

Contribution

It presents a novel application of vibrational photothermal microscopy to visualize and quantify thermophoretic effects inside cells, including the discovery of reversed transport directions.

Findings

Reversed molecular transport in cytoplasm and nucleus.

Reduced thermophoretic activity under CO2-depleted conditions.

Observation of glass formation during cell death.

Abstract

Vibrational microscopy provides label-free, bond-selective chemical contrast by detecting molecular vibrations, making it invaluable for biomedical research. While conventional methods rely on the direct detection of Raman scattering or infrared absorption, recently developed vibrational photothermal (ViP) microscopy achieves chemical contrast indirectly through refractive index (RI) changes. This indirect approach enables unique imaging capabilities beyond traditional chemical imaging. Here, we introduce a novel application of ViP microscopy: label-free intracellular thermophoretic (Soret) imaging, which visualizes biomolecular transport driven by temperature gradients. ViP-induced Soret (ViPS) imaging leverages a steady-state temperature distribution generated by optical heating through vibrational photothermal effect, combined with time-resolved RI imaging via optical diffraction tomography (ODT). Using ViPS imaging, we measured thermophoretic behavior in living COS7 cells, determining intracellular diffusion and Soret coefficients. Notably, we observed a reversed direction of molecular transport (negative Soret effect) in the cytoplasm compared to the nucleus, possibly driven by thermophoresis-induced diffusiophoresis. Furthermore, time-lapse imaging under CO2-depleted conditions revealed a remarkable reduction in thermophoretic activity, suggesting glass formation during the dying process, likely due to polymer aggregation. ViPS imaging represents a new frontier in intracellular thermophoretic studies, expanding the capabilities of vibrational microscopy.