Toward the Next Frontiers of Vibrational Bioimaging

Abstract

Chemical imaging based on vibrational contrasts can extract molecular information entangled in complex biological systems. To this end, nonlinear Raman scattering microscopy, mid-infrared photothermal (MIP) microscopy, and atomic force microscopy (AFM)-based force-detected photothermal microscopies are emerging with better chemical sensitivity, molecular specificity, and spatial resolution than conventional vibrational methods. Their utilization in bioimaging applications has provided biological knowledge in unprecedented detail. This Perspective outlines key methodological developments, bioimaging applications, and recent technical innovations of the three techniques. Representative biological demonstrations are also highlighted to exemplify the unique advantages of obtaining vibrational contrasts. With years of effort, these three methods compose an expanding vibrational bioimaging toolbox to tackle specific bioimaging needs, benefiting many biological investigations with rich information in both label-free and labeling manners. Each technique will be discussed and compared in the outlook, leading to possible future directions to accommodate growing needs in vibrational bioimaging.

The Duality of Raman Scattering

“…Since the advent of SRS microscopy, it has launched a rapidly growing field of chemical imaging. By leveraging advances in laser spectroscopy, imaging probes, and data science, SRS microscopy can map the distribution of chemical bonds in three-dimensional samples with high speed, specificity, and resolution, making a broad impact in chemistry, life science, and material science. ,,− However, the theory underlying SRS seems to lag behind the experiments. Indeed, the discovery of the SRS effect in 1962 was serendipitous .…”

Nitrile Vibrational Lifetimes as Probes of Local Electric Fields

“…However, the potential of electric field mapping in cells with nitrile vibrations has not yet been fully realized, mainly due to the limited sensitivity of linear IR absorption spectroscopy . To improve the sensitivity of vibrational bioimaging, a handful of ultrasensitive vibrational microscopy techniques have been developed. − We recently reported one such technique, termed bond-selective fluorescence-detected IR-excited (BonFIRE) spectro-microscopy (Figure ). BonFIRE is a picosecond mid-IR (MIR)–near-IR (NIR) double-resonance fluorescence technique (Figure b) with applicability to a wide range of fluorescent dyes and excellent biocompatibility.…”

Atomic Force Microscopy-Based Nanoscale Infrared Techniques for Catalysis

“…A straightforward approach is to increase the power density of the utilized IR laser. The past five years have witnessed rapid advancements in optical parametric oscillator (OPO)-based difference frequency generation (DFG) sources, optical parametric amplifiers (OPAs), and quantum cascade lasers (QCLs). , These advancements have enabled the attainment of wider spectral ranges, higher repetition rates, and greater pulse energies, facilitating their extensive use in AFM-based nano-IR techniques for generating detectable signals of molecular monolayer-level samples. Additionally, robust synchrotron-based IR beams covering the entire mid-IR range of 500–5000 cm –1 are also ideal for this purpose .…”