Molecular component distribution imaging of living cells by multivariate curve resolution analysis of space-resolved Raman spectra

Abstract

Abstract. Label-free Raman microspectroscopy combined with a multivariate curve resolution (MCR) analysis can be a powerful tool for studying a wide range of biomedical molecular systems. The MCR with the alternating least squares (MCR-ALS) technique, which retrieves the pure component spectra from complicatedly overlapped spectra, has been successfully applied to in vivo and molecular-level analysis of living cells. The principles of the MCR-ALS analysis are reviewed with a model system of titanium oxide crystal polymorphs, followed by two examples of in vivo Raman imaging studies of living yeast cells, fission yeast, and budding yeast. Due to the non-negative matrix factorization algorithm used in the MCR-ALS analysis, the spectral information derived from this technique is just ready for physical and/or chemical interpretations. The corresponding concentration profiles provide the molecular component distribution images (MCDIs) that are vitally important for elucidating life at the molecular level, as stated by Schroedinger in his famous book, "What is life?" Without any a priori knowledge about spectral profiles, timeand space-resolved Raman measurements of a dividing fission yeast cell with the MCR-ALS elucidate the dynamic changes of major cellular components (lipids, proteins, and polysaccharides) during the cell cycle. The MCR-ALS technique also resolves broadly overlapped OH stretch Raman bands of water, clearly indicating the existence of organelle-specific water structures in a living budding yeast cell.

Characterization of different substrates for Raman spectroscopic imaging of eukaryotic cells

“…[1][2][3][4][5][6][7][8][9][10][11][12] In vitro measurement requires placing the biological specimen on a substrate as a sample holder. However, the Raman spectra of the sample holder itself need to be dormant so that no Raman spectral signature of the holder itself is interfering with the sample's Raman signals.…”

Parallelized shifted‐excitation Raman difference spectroscopy for fluorescence rejection in a temporary varying system

“…MCR‐ALS is an alternative method to PCA in finding bilinear matrix decomposition of the input data matrix. By the use of non‐negativity constraint, which reflects the physical nature of Raman scattering, the method tends to find a chemically interpretable decomposition solution . The user must supply the number of components, and in the present case, we designated two components based on the PCA result.…”

Raman spectroscopic study on the excystation process in a single unicellular organism amoeba (Acanthamoeba polyphaga)

“…During the excystation processes, the transportation mediators play a very important role for transformation from a dormant/resting to an active metabolic state. The spectral bands were assigned according to the literature; 19,20,[22][23][24] characteristic Raman bands for living cells were observed. 1, the amoeba images during the encystation and excystation are presented.…”

“…The non-invasive advantage renders the possibility for investigation in living samples. 19,20 In this work, we present the Raman spectral analysis of amoeba (Acanthamoeba polyphaga) in the cyst stages. [11][12][13][14] New methods based on Raman spectroscopy (such as surface-enhanced Raman scattering, coherent anti-stokes Raman scattering, Raman mapping, etc.…”