Multimodal vibrational imaging of cells

Fourier Transform Infrared (FTIR) and Raman imaging offer complementary information about the spatial location of molecules within cells. In this paper we investigate the integration of both imaging modalities in an extended image containing unique FTIR and Raman spectra for each pixel. Two types of cells were investigated: red blood cells infected with the Plasmodium falciparum parasite and Micrasterias, a desmid microalgal species. The microscope configuration and pixel size were selected specifically for acquiring images with the same pixel size, and samples were fixed to a sample holder in order to measure the same cells with the same orientation. Images were first analysed individually and then registered in an extended matrix containing an FTIR and a Raman spectra for each pixel. The results indicated that the combination of both techniques provide complementary information not evident in the analysis of individual images. The assignment of haemozoin FTIR bands from malaria trophozoites was only possible after correlating the FTIR spectrum with the Raman spectrum. The correlation of the Raman and FTIR spectral variables using statistical heterospectroscopy (SHY) enabled the assignment of overlapping lipid and carbohydrate bands. In summary, although the approach can be computing intensive the complementarity of the two techniques in terms of pixel resolution, signal to noise ratio and the assignment of vibrational modes makes this a powerful approach to diagnostic imaging.