Localizing and Identifying Living Bacteria in an Abiotic Environment by a Combination of Raman and Fluorescence Microscopy

摘要

A fast, easy, and reliable identification of microorganisms is indispensable in many fields such as medicine, food production, or the pharmaceutical industry. However, in native samples, biotic particles often appear together with abiotic particles. Therefore, it is a prerequisite that biotic particles can be differentiated from abiotic particles appearing in the identification setup. In addition, for many applications, not all microorganisms are of interest but only the living ones. Therefore, in this contribution, different bacteria species were stained with a live/dead staining kit (SYTO 9 and propidium iodide) prior to Raman spectroscopic identification. Since only living and dead microorganisms are getting stained by SYTO 9 or PI, biotic particles can easily be spotted and localized in-between abiotic particles. By using a Raman laser excitation wavelength outside the absorption band of the dye, fluorescence-free Raman spectra were obtained. The living cells were identified by means of Raman spectroscopy in combination with a support vector machine. Furthermore, the localization of bacterial cells in a mix of abiotic particles is demonstrated.