Fourier transform mid-infrared imaging and rapid evaporative ionization mass spectrometry imaging in FFPE colorectal...

Hungary ranks highest in the world regarding both the incidence and death rates of colorectal cancer. Novel analytical methods have surfaced in the past decade and are infiltrating the field of medicine. Infrared spectroscopy (IR) and mass spectrometry (MS) are frequently used for such measurements; however, multimodal techniques resulting from the combination of these have not yet gained widespread adoption.

Tissue microarray (TMA) of colorectal adenocarcinomas, liver metastases, and surrounding colon tissues were prepared at the Institute of Pathology, Forensic, and Insurance Medicine, Semmelweis University. From the block, 2‑ and 10-µm sections were cut for infrared and MS imaging, respectively. Infrared measurements were performed with a Perkin Elmer Spotlight microscope (Perkin Elmer Inc., Waltham, MA, USA) on samples with an area of 2200 × 2200 µm², in the wavenumber range of 4000-750 cm^-1. For the mass spectrometry measurements, a custom Waters Xevo G2 XS QToF-type lipid group focusing device (Waters Corporation, Santa Barbara, CA, USA) was used on samples with an area of 10 mm × 10 mm between 150-900 m/z values. The uniqueness of the device comes from the fact that a high-energy laser had been added for the purpose of ionizing the sample tissue.

The spectral results from the samples were noise filtered in both methods to separate the sample signal from the background. Overall, 41 cores were suitable for further data processing. For the measurements with infrared spectroscopy, the three-class classification resulted in accuracy of 0.71 and 0.71 using C‑support vector classification (SVC) and eXtreme Gradient Boosting (XGBoost) algorithms. For the analysis of the mass spectrometry data, 0.65 and 0.70 accuracy values were observed.

Identical TMA slides have been imaged with two analytical methods. The performance of the separating algorithms was very close, which underscores the efficacy of the spectroscopy and spectrometry approach. A vector database of the spectra could be combined with the whole-slide image pixels obtained by a traditional digital scanner. A further step could be a thickness-optimized measurement, which would determine the setting for the combined measurement of the exact same sample.