How Infrared Microscopy Works & How We Save You Time & Money

FTIR spectroscopy is a well-established technique used to analyze a substance’s makeup by analyzing a material’s IR absorption and emission characteristics. The Fourier transform of FTIR refers to a specific mathematical principle that takes raw data and translates it into its corresponding spectrum.

When combined with FTIR microscopy, researchers can visualize a sample as the spectrometer analyzes it.

The FTIR spectrometer is used for spectral analysis while the detector works to image the sample. When this imaging occurs, it defines specific sample areas, or regions, as pixels. The spectrometer analyzes each pixel’s infrared spectra.

Furthermore, the microscope allows for the analysis of much smaller samples than FTIR spectroscopy alone. Nonetheless, the sample size can vary from one FTIR microscope to another. Other feature differences include fully- vs. semi-automated analysis/visualization and the ability to capture and store images or videos of the samples as the analysis occurs.

FTIR imaging is ideal for research related to materials science, failure analysis, particle analysis, and chemical analysis.

This measurement method is a simple one and requires little sample prep. However, it has its limitations, like reflection measurements that are somewhat limited to certain types of samples. One such sample, according to Shimadzu, is foreign matter adhering to a highly reflective metal substrate.

The reflection technique is performed using a Cassegrain objective, which differs from a traditional microscope’s objective lens. The Cassegrain objective uses a primary and secondary mirror to send the IR beam onto the sample at a specific degree.

While simple in design, reflection measurement does not work well with dark samples. IR reflective materials work best with this objective and technique.

Like the reflection sample technique, attenuated total reflectance (ATR) is simple and does not require extensive sample preparation. It uses specialized crystal materials to obtain a sample’s chemical information.

More specifically, it’s used to measure the changes in an internally reflected IR beam when the beam comes in contact with a sample. The beam is directed onto an optically dense crystal with a high refractive index at a certain angle, which creates an evanescent wave that extends beyond the surface of the crystal into the sample held in contact with the crystal.

The evanescent wave becomes attenuated in regions of the IR spectra where the sample absorbs energy. This attenuated beam then returns to the crystal and exits the opposite end, directed to the detector in the IR spectrometer. The detector records the attenuated IR beam as an interferogram signal which is used to generate an IR spectrum.

Transmission measurement is widely considered the most traditional method of analyzing solids, liquids, and gases. This analysis provides high sensitivity and detection of infrared sampling techniques. When performing FTIR analysis, the research must place the sample into the infrared beam of the spectrometer.

Two Cassegrain objectives are employed to measure transmission: a focusing lens and a condenser lens. The focusing objective works in the same fashion as a reflection objective. Light passes through the sample at the focal point and then hits the condenser, which collimates the beam into the detector.

Samples must be thin (the recommended size is less than 50 microns) and must be prepared into a pellet, mull, or film. Using this sampling technique, excellent spectra can be obtained for many samples. This includes various polymers, such as soluble polymers, thin polymer films, non-carbon-filled dark polymer films, irregular-shaped polymers, and regular-shaped polymers. This also includes organic powders in pellet or mull form and thermoplastic powders.

There are various ways to generate an IR image. The simplest is to perform single measurements with defined distances on a sample and is referred to as single point mapping.

That said, creating IR images more effectively requires specialized detectors, which can visualize the entire IR spectrum. One of the most common detectors used in IR microscopy is the focal-plane array (FPA). This image sensor employs a variety of light-sensing pixels at the microscope’s focal plane.

The detector works by picking up photos of specific wavelengths and then creating an electric charge proportional to how many photons it detects. This electric charge is then analyzed and used to construct an image. This specific type of detector can gather up to 16,000 pixels simultaneously, which allows for IR imaging.

IR microscopy is a powerful imaging tool for scientists and will continue to be so into the future. However, it can take a whole day to analyze the data gathered. This makes IR microscopy reasonably limited in its usefulness in the clinical setting. However, a team of researchers at the Ruhr-Universität Bochum (RUB) simplified this measurement step and reduced the analysis time from a day to a few minutes.

They achieved this by using quantum cascade lasers (QCLs). Unlike conventional IR microscopes, QCL-based IR microscopes can employ only a single frequency that cuts down on the measuring and analyzing phases. These QCL-based IR can be used in the label-free classification of cancer tissues.

In a study of 110 tissue samples, this method proved to have 96% sensitivity and 100% specificity compared to histopathology, the gold standard for cancer diagnosis. A rapid cancer diagnosis is vital in cancer treatment. The earlier doctors can diagnose it, the better chances a patient will have positive treatment options.