Multidimensional Information on the Chemical Composition of Single Bacterial Cells by Confocal Raman Microspectroscopy

Abstract

In many biotechnological processes, living microorganisms are used as biocatalysts. Biochemical engineering science is becoming more aware that individual cells of an organism in a process can be fairly inhomogeneous regarding their properties and physiological status. Raman microspectroscopy is a novel approach to characterize such differentiated populations. Cells of the anaerobic bacterium Clostridium beijerinckii were dried on transparent support surfaces. The laser beam of a confocal Raman microscope was focused on individual cells viewed through the objective. Single bacterial cells in size approximately 1 microm and sample mass approximately 1 pg could be analyzed within a few minutes, when placed on a calcium fluoride support and using excitation at 632.8 nm. Spectral features could be attributed to all major cell components. Cells from a morphologically differentiated culture sample showed different compositions, indicating the presence of subpopulations. As a reference, the storage polymer granulose was detected. The multidimensional information in Raman spectra gives a global view on all major components of the cell at once, complementing other more specific information-rich methods for single-cell analysis. The method can be used, for example, to study heterogeneities in a microbial population.