## 1 Introduction  
In recent years, vibrational spectroscopic techniques, such as Raman Microspectroscopy (RMS) and Fourier Transform Infrared (FTIR) spectroscopy, have been employed as emerging tools for studies in the biomedical field, including those on the structural and macromolecular composition of tissues and cells [1, 2, 3]. Both of these non-destructive and label-free analytical techniques, based on the interaction between matter and electromagnetic radiation, provide relevant information on the chemical bonds, and hence the functional groups of the macromolecules within the analyzed sample [4, 5]. RMS is defined as a scattering technique; indeed, it measures the two-photon inelastic light scattering produced when the photons from monochromatic electromagnetic radiation interacting with matter are scattered from the samples with an energy higher or lower in respect to the incident one [6, 7, 8]. It does not require extrinsic contrast-enhancing agents and allows one to obtain reproducible and reliable results on the chemical composition of the sample, with minimal to no preparation [8, 9]. On the other hand, FTIR is based on the absorption of electromagnetic radiation in the IR range; hence, it is an absorption spectroscopy [8, 10]. Infrared spectroscopy can be applied in transmission mode for samples transparent to the radiation, or in reflectance mode in the case of solid and non-transparent materials [10]. In this latter case, IR spectra are collected using Attenuated Total Reflectance-Fourier transform InfraRed spectroscopy (ATR-FTIR), a contact sampling method that exploits a crystal with a high refractive index; this technique is based on the analysis of the 'evanescent wave', originating from the crystal and penetrating within the material for a fraction of its wavelength [11; 12]. Infrared and Raman techniques also differ in the measured molecular vibrations, with the first detecting changes in the dipole moment, and the second detecting variations in the polarizability of the molecule [13]. This difference means that Raman spectroscopy can detect the symmetric vibrations of non-polar groups, while IR spectroscopy better measures the asymmetric vibrations of polar groups [8]. Therefore, RMS and FTIR are completely complementary techniques; the different physical phenomena they rely on determine differences in terms of provided information and applicability; hence, each of them show specific advantages and disadvantages, also in their application to biological systems [4; 10]. For example, IR spectra are not affected by fluorescence, which often influences Raman measurements [14]. On the other hand, RMS is more suitable for the analysis of samples in aqueous solution; in fact, Raman spectra are not altered by water, which conversely massively influences IR spectra and requires complex subtraction procedures [2; 14]. In the last decades, Raman and FTIR instrumentations have been significantly improved by their coupling with optical microscopes that let perform the imaging analysis. In this way, by mapping micro-areas of interest, it is possible to combine the information derived from the morphological analysis of the sample with its punctual chemical composition [15; 16]. However, given the different physical phenomena on which the two techniques rely, IR spectroscopy requires 5-10 \(\upmu\)m-thick tissue sections for analysis in transmission mode [9; 17; 18], while RMS can be applied to solid and thicker samples. Moreover, since RMS is based on light scattering, it can be exploited in a confocal mode by focusing the laser not only on the surface of the sample, but also at a specific depth [7; 19]. In addition, RMS provides maps with a better spatial resolution than FTIR, allowing the detection of biological modifications which cannot be highlighted using more traditional techniques [19].