Influence of measurement mode on the results of glioblastoma multiforme analysis with the FTIR microspectroscopy

Fourier Transform Infrared (FTIR) microspectroscopy is well known for its effectiveness in spectral and biochemical analyses of various materials. It enables to determine the sample biochemical composition by assigning detected frequencies, characteristic for functional groups of main biological macromolecules. In analysis of tissue sections one of two measurement modes, namely transmission and transflection, is usually applied. The first one has relatively straightforward geometry, hence it is considered to be more precise and accurate. However, IR-transparent media are very fragile and expensive. Transflection does not require expensive substrates, but is more prone to disruptive influence of Mie scattering as well as electric field standing wave effect. The excessive comparison of spectra' characteristics, obtained via both measurement modes, was performed in this paper. By the means of Mann-Whitney non-parametrical U test and PCA, the comparison of results obtained with both modes and assessment of usefulness of IR spectra obtained with transmission and transflection modes to differentiate between healthy and GBM-affected tissue, were performed. The main objective of the presented research is to compare the results of FTIR analysis of unfixed biological samples performed with transflection and transmission mode. In frame of the study we demonstrated the discrepancies between results of biochemical analysis performed based on data obtained with transmission and transflection. Such observation suggests that caution should be taken in drawing conclusions from the results obtained with transflection geometry, as its more prone to disruptive effects. Despite that, IR spectra developed with both modes allowed to distinguish GBM area from healthy tissue, which proves their diagnostic potential. Especially, application of the ME-EMSC correction of spectra before PCA enhances the performance of both methods to distinguish the analysed tissue areas.