Enhancement of the super-resolved optical photothermal infrared signal at lower temperatures

Optical photothermal infrared (O-PTIR) microspectroscopy is an emerging label-free technique that achieves infrared super-resolution and has found widespread applications, primarily in soft materials. While the photothermal effect underlying O-PTIR has been extensively studied at room temperature, there was an open question whether the signal itself would benefit from a lower temperature, as the photoinduced heating could be more localized, thus increasing the signal. In this work, we integrated a temperature-controlled stage into an O-PTIR microscope, enabling the first spectroscopic measurements and imaging of samples at subzero temperatures down to -100 °C. Using tristearin, biological tissue, and polymer samples, we demonstrated that cooling significantly enhances the O-PTIR signal up to three times, with a maximum observed in the -20 to -40 °C range. This enhancement leads to an improved analytical sensitivity, reducing the limit of detection for a thin protein film from ca. 400 nm to ca. 300 nm. We further discuss the underlying physical mechanism with theoretical modeling, including thermal conductivity and heat dissipation effects, that contribute to the observed temperature-dependent signal amplification. Overall, this study establishes temperature-controlled O-PTIR as a powerful new approach for high-sensitivity, high-resolution infrared analysis, thereby expanding its applicability to a broader range of materials and experimental conditions.