Nano-IR Applications in Life Science

Infrared nanospectroscopy (nano-IR) integrates the chemical specificity of infrared absorption with nanoscale spatial resolution, enabling vibrational spectroscopic analysis well below the diffraction limit. By combining atomic force microscopy (AFM) with tunable infrared illumination, nano-IR probes local absorption through photothermal expansion or near-field scattering, yielding chemical maps with <20 nm of spatial resolution. This capability uniquely addresses the hierarchical complexity of biological matter—from biomolecular assemblies to sub-cellular structures—in situ and label-free. In life sciences, nano-IR has rapidly expanded beyond materials research to tackle fundamental and applied biological questions. Key applications include: Protein and peptide structural characterization [1-10], where nano-IR resolves secondary structure variations in amyloid fibrils and aggregates with spatial specificity unattainable by conventional IR. Cellular biochemistry [11-19] through sub-cellular mapping of lipids, proteins, nucleic acids, and metabolites, enabling correlation of chemical composition with morphology in single cells. Recent developments extend nano-IR [19-21] into liquid environments critical for maintaining native biological function. We highlight the design and implementation of microfluidic cell platforms compatible with nano-IR measurement, addressing challenges in optical coupling, fluid handling, and thermal management. We employ a membrane-based liquid cell architecture coupled with scattering-type scanning near-field optical microscopy (s-SNOM). A critical element of this design is a 10-nm-thin silicon nitride (SiN) membrane that physically separates the liquid sample compartment from the AFM tip and optical components (Fig. 1). The ultrathin SiN layer serves several key functions: it prevents evaporation of the liquid, protects the tip from contamination, and enables mechanically stable near-field measurements over extended periods without realignment of the optics, even when the sample is exchanged. Importantly, the SiN membrane is thin enough to allow the optical near field generated at the tip apex to penetrate through into the liquid and interact with submerged nanoscale objects, making label-free, chemically specific measurements in native aqueous conditions feasible. Nano-IR’s unique combination of nanoscale spatial resolution, chemical specificity, and compatibility with liquid environments positions it as a transformative tool across life science applications. The transient nanoscopy of single lipid vesicles in water demonstrates that dynamic, label-free vibrational spectroscopy at biologically relevant time and length scales is achievable. Coupling nano-IR with engineered microfluidic liquid cells further expands opportunities for in situ studies of cells, biomolecular assemblies, and biochemical interactions under native conditions. Design and principle of the microfluidic cell. The AFM tip performs operation in air, tapping over a surface of SiN membrane of ∼10 nm thickness. s-SNOM method has a penetration depth of <100 nm, allowing subsurface medium chemical analysis, also being sensitive to specimens, located in aquatic phase.