Mid-Infrared Photothermal Microscopy: Principle, Instrumentation, and Applications

Abstract

Midinfrared photothermal (MIP) microscopy, also called optical photothermal infrared (O-PTIR) microscopy, is an emerging tool for bond-selective chemical imaging of living biological and material samples. In MIP microscopy, a visible probe beam detects the photothermal-based contrast induced by a vibrational absorption. With submicron spatial resolution, high spectral fidelity, and reduced water absorption background, MIP microscopy has overcome the limitations in infrared chemical imaging methods. In this review, we summarize the basic principle of MIP microscopy, the different origins of MIP contrasts, and recent technology development that pushed the resolution, speed, and sensitivity of MIP imaging to a new stage. We further emphasize its broad applications in life science and material characterization, and provide a perspective of future technical advances.

Mapping Enzyme Activity in Living Systems by Real-Time Mid-Infrared Photothermal Imaging of Nitrile Chameleons

“…The recently developed MIP microscope enables infrared absorption chemical imaging with submicron spatial resolution by sensing the photothermal effect induced by mid-infrared absorption with a visible light. 23,24 Importantly, such indirect measurement mitigates the huge water absorption issue and allows mid-infrared imaging in aqueous environment with high contrast. 25 The reported samplescan MIP microscope needs a pixel dwell time ranging from 500 μs to 10 ms, 23,24 making it difficult to resolve the dynamics of living systems.…”

“…23,24 Importantly, such indirect measurement mitigates the huge water absorption issue and allows mid-infrared imaging in aqueous environment with high contrast. 25 The reported samplescan MIP microscope needs a pixel dwell time ranging from 500 μs to 10 ms, 23,24 making it difficult to resolve the dynamics of living systems. To improve the MIP imaging speed, widefield approach using camera as the sensor has been developed for spatially multiplexed detection.…”

“…While label-free MIP microscopy succeeds in visualizing the abundant cellular components (e. g. proteins and lipid droplets), 23,24 direct observation of a specific enzymatic reaction remains difficult because the native MIP signals from products are likely immersed into the strong background generated by other biomolecules. To address this challenge, we synthesized a kind of nitriletagged bioorthogonal enzyme activity probes.…”

High-Throughput Antimicrobial Susceptibility Testing of Escherichia coli by Wide-Field Mid-Infrared Photothermal Imaging of Protein Synthesis

“…Mid-infrared photothermal (MIP) microscopy was recently developed to overcome the abovementioned limitations. In MIP imaging, visible light detects the photothermal effect induced by IR absorption. − The sub-micrometer spatial resolution and high sensitivity of MIP imaging have been demonstrated in the detection of single viruses, bacteria, − cells, ,,,− and tissues. , The direct imaging of lipid metabolism in living cells has been shown by combining MIP with isotopic labeled fatty acids . The majority of the developed MIP systems are based on point-scan measurement, where both the IR pump and visible probe are focused and an image is created from stage-scanning.…”

Exploration of utility of combined optical photothermal infrared and Raman imaging for investigating the chemical composition of microcalcifications in breast cancer

“…38,39 More recently the technique of optical photothermal infrared (O-PTIR) spectroscopy has been developed, which measures the photothermal response of the sample to pulses of single frequency mid-IR radiation. 40 The thermoelastic expansion following the resonant absorption leads to a change in the local refractive index of the sample. The photothermal response is detected by a non-destructive visible or NIR laser probe which determines the spatial resolution of the measurement.…”