Super-resolution mid-infrared spectro-microscopy of biological applications through tapping mode and peak force tapping mode atomic force microscope

[Display omitted] •Near-field optical spectro-microscopic methods based on the tapping mode and peak force tapping AFM operation can bypass the diffraction limit and offer superb surface sensitivity due to tip-enhancements.•s-SNOM has been extensively utilized in chemical identification and nanoscale heterogeneity imaging in studying plant biology, biomolecule imaging, cell and tissue biology, and biomaterial engineering.•Relatively young PiFM and PFIR microscopy has been utilized to study a range of biological specimens.•The development of s-SNOM, PiFM and PFIR microscopy in water will enable visualizing spatial–temporal distribution of chemical compositions in biological systems’ native state.•Further technical and methodological development of s-SNOM, PiFM and PFIR microscopy promises a wealth of opportunities in in situ biological and pharmaceutical studies. Small biomolecules at the subcellular level are building blocks for the manifestation of complex biological activities. However, non-intrusive in situ investigation of biological systems has been long daunted by the low spatial resolution and poor sensitivity of conventional light microscopies. Traditional infrared (IR) spectro-microscopy can enable label-free visualization of chemical bonds without extrinsic labeling but is still bound by Abbe’s diffraction limit. This review article introduces a way to bypass the optical diffraction limit and improve the sensitivity for mid-IR methods – using tip-enhanced light nearfield in atomic force microscopy (AFM) operated in tapping and peak force tapping modes. Working principles of well-established scattering-type scanning near-field optical microscopy (s-SNOM) and two relatively new techniques, namely, photo-induced force microscopy (PiFM) and peak force infrared (PFIR) microscopy, will be briefly presented. With ∼ 10–20 nm spatial resolution and monolayer sensitivity, their recent applications in revealing nanoscale chemical heterogeneities in a wide range of biological systems, including biomolecules, cells, tissues, and biomaterials, will be reviewed and discussed. We also envision several future improvements of AFM-based tapping and peak force tapping mode nano-IR methods that permit them to better serve as a versatile platform for uncovering biological mechanisms at the fundamental level.