A Spectrum of Capabilities in FTIR Microspectroscopy

Frederick William Herschel (1738–1822) composed 24 symphonies. He also discovered Uranus, the first planet ever detected with equipment more powerful than human eyesight, although he assumed all celestial bodies to be populated, including the interior of the Sun. His dedication to the study of reflected light spurred prodigious innovations in astral telescopes and laid bare the raw power of Newton’s mathematical laws governing prismatic dispersion. Herschel inferred infrared (IR) radiation from his observations of temperature changes in samples subjected to sunlight outside the visible spectrum. He concluded that the absorption of IR energy was proportional to the temperature change observed as a sample moved through different wavelengths. As luck would have it, molecular vibrations occur at the frequencies that correspond to IR wavelengths. When radiant energy matches the energy of a particular vibration caused by stretching and bending along chemical bonds, the resulting spectrum is equivalent to a molecular fingerprint. The field of spectroscopy emerged from these relationships, and systems that were later adopted widely among chemists and technicians as quality control tools have their origins in bedrock observations and theories regarding the nature of electromagnetic radiation and hence the fundamental properties of space and time. The special theory of relativity, for example, relies upon the Lorentz factor, a mathematical caveat to accommodate the result of the Michelson– Morley experiment. In this seminal experiment, measurements of reflected light split by a silvered mirror in a device called a Michelson interferometer unexpectedly negated the fin-de-siècle scientific dogma that Earth moved through a luminous ether that influenced the speed of light.

Herschel’s son John developed the first IR images, and IR spectra were soon used to characterize chemical compounds. However, the technology was initially limited to using dispersive techniques, in which radiation of known individual frequencies is used to bombard targets, and spectra are collected wavelength by wavelength. Dispersive spectroscopy is still frequently used, for instance, to characterize spectra obtained via X-ray backscatter that results as a by-product of other techniques, such as electron microscopy. Like so many other leaps forward, the propulsive force of wartime annihilation helped catalyze improvements in IR spectroscopy. In World War II, some missile guidance systems used diffraction of IR radiation through crystallized salts; similar strategies were then adapted in miniature to facilitate the first commercial spectroscopy devices. Incorporation of the Michelson interferometer vastly increased throughput by enabling simultaneous bombardment with a range of frequencies via a constantly moving mirror to create a variable time delay. The result, an interferogram, must then be reconstituted into corresponding absorption spectra using Fourier transform analysis to plot transmittance against wavenumber.

Fourier transform infrared spectroscopy (FTIR) has long been a valuable tool to facilitate confirmation of molecular structure through analysis of spectra in comparison to reference spectra. It is frequently used to identify contaminants in organic chemistry synthesis and pharmaceutical production chains. FTIR has several advantages over dispersive techniques, including the ability to obtain more comprehensive spectra and to do so more quickly. Additionally, the energy imparted to the sample via continuous reflection, rather than stepwise grated dispersion, allows for higher sensitivity and therefore a greater signal-to-noise ratio. Intrinsically, the diffraction limit of IR radiation defines the resolution of molecular analysis; addition of various microscopy platforms to FTIR equipment has enabled transcendence of previous limits and allowed, at minimum, higherorder applications such as surface mapping of conformational changes during drug-target binding. Consequently, FTIR has moved from being a quality control tool to a powerful analytical and predictive tool.

Beyond quality control: applications in nanomaterials and medicine

One field in which FTIR has made great inroads is the production and analysis of nanomaterials. The word “nanomaterials” usually elicits a vague and static concept of something very small. However, a partial list of nanomaterial classes includes quantum dots, nanospheres, nanolenses, nanorods, nanofibers, nanocubes, nanosheets, and nano-onions. Each of these has a distinctive topographical footprint in zero to three dimensions within and beyond nano-space. Researchers can use FTIR microspectroscopy to characterize properties and irregularities both of nanomaterials and of macromaterials at the nanoscale level. Consequently, they can make structure– function relationship predictions and strategize improvements. For instance, in computer engineering, detailed information about nanomaterial structure can guide efforts to improve highcapacity storage and high-resolution imaging. In medicine, FTIR microscopy can be used at the nanoscale to improve targeting specificity or lower toxicity in drug delivery systems.

Constantly evolving FTIR platforms adapted to novel microscopic lenses, computational algorithms, and precision technologies, such as quantum cascade laser imaging, have the potential to demolish established ceilings in the speed and accuracy of medical diagnostics. False-negative and -positive histopathology results from subjective analysis of stained biopsies could conceivably be eliminated by objective FTIRbased digital standards. Cancer prognoses and treatments could acquire greater accuracy through quantitative FTIR-based mapping of tumor boundaries and real-time intracellular analysis of drug-target interactions. Label-free detection of cell-surface disease markers without addition of fluorophores and chemical surrogates that can confound analysis promises to streamline and optimize clinical diagnostics. Moreover, FTIR can potentially define novel disease markers by sorting and analyzing individual extracellular vesicles according to their distinct biochemical and optical properties.

User-friendly FTIR microscopy

The origins of FTIR in observations, calculations, and theorems by great minds concerned with the nature and origin of the universe begs the question of whether a garden-variety scientist can obtain and analyze data without first obtaining at least an extra doctoral degree. Commercially, providers have optimized automation and software interfaces within FTIR microscopy equipment to be used with minimal training. A decreased footprint also makes it easier to bring the technology into individual laboratories. Bruker offers the LUMOS as an entry-level model with a broad range of capabilities for particle identification, chemical composition analysis, and measurements of defects and contaminants. Its HYPERION series is more suitable for dedicated materials R&D labs or core facilities, with major upgrades in frequency detection range and objectives arranged to accommodate different types of samples. Analogously, Thermo Nicolet FTIR devices, which run the gamut in capability and resolution from the iN5 to the Continuμm, are configured for straightforward quality control studies. With the products available, there is a spectrum of capabilities in FTIR microspectroscopy.

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