Mapping of single cells by near infrared Raman microspectroscopy

Abstract

Objective of our work is the development of new diagnostic methods for detection of tissue states (e.g. tumors, necrosis) in vivo. Therefore, it is important to understand the spectral features of pure individual components of tissues, that means cells and subcellular components. Raman spectroscopy has promising potential as an analytical tool for clinical applications because it can probe the chemical composition and molecular structure of such complex systems. Furthermore, the spatial resolution of Raman microspectroscopy in the low micrometer scale and its ability to probe samples under in vivo conditions allow new insights into living single cells without the need for fixatives, markers or stains. In a first set of experiments we prepared human embryonic lung epithelial fibroblasts, human osteogenic sarcoma cells and human astrocytoma cells. Raman mapping data sets were acquired with 1 μm step size and 1 min exposure time per spectrum using 785 nm excitation wavelength. Principal component analysis (PCA) was used for evaluation of the spectral maps. Bands in individual spectra were assigned to proteins, lipids, cholesterol and nucleic acids. Based on this information, the main cellular constituents of freeze dried cells and living cells in media were identified in score plots of principal components.