Multi-contrast wide-field mid-infrared photothermal imaging

Abstract: Wide-field mid-infrared photothermal (MIP) imaging enables high-speed, chemically specific imaging at sub-micrometer spatial resolution through several possible contrast mechanisms, including scattering change, fluorescence modulation, and phase change. Each mechanism offers distinct advantages, yet existing wide-field MIP instruments have typically been limited to a single contrast mechanism so far, requiring a dedicated instrument for each. Here, we introduce a switchable multi-contrast wide-field MIP microscope that allows the photothermal readout mechanism (based on scattering-, fluorescence-, or quantitative phase imaging (QPI)) to be matched to the sample and imaging conditions. We characterize the spectral fidelity and signal-to-noise (SNR) ratio of each mechanism using 5 μm polystyrene (PS) beads, and the effective spatial resolution using 300 nm PS beads. Scattering- and fluorescence-based detection achieved a sub-500 nm measured point spread function full width at half maximum, while QPI-based detection reached approximately 900 nm but provided up to 26-fold higher SNR than scattering. We further demonstrate imaging across a range of samples, including A549 cells in buffer, mouse lung tissue section, bacteria, and SU8 polymer at 1.4 and 5 μm thickness, with a field of view exceeding 200 μm. Comparison across these samples shows that each contrast mechanism offers distinct trade-offs in resolution, sensitivity, and applicability, guiding readout-strategy selection by sample type. These results establish a flexible, switchable multi-contrast platform for wide-field MIP imaging, with broad applicability across polymer and biological samples.