Fluorescence Imaging of Mycobacterial Infection in Live Mice Using Fluorescent Protein-Expressing Strains.
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Fluorescence imaging has been applied to various areas of biological research, including studies of physiological, neurological, oncological, cell biological, molecular, developmental, immunological, and infectious processes. In this chapter, we describe methods of fluorescent imaging applied to examination of subcutaneous and pulmonary mycobacterial infections in an animal model. Since slow growth of Mycobacterium tuberculosis (Mtb) hinders development of new diagnostics, therapeutics, and vaccines for tuberculosis (TB), we developed fluorescent protein (FP) expressing mycobacterial strains for in vivo imaging, which can be used to track bacterial location and to quantitate bacterial load directly in living animals. After comparison of imaging data using strains expressing different fluorescent proteins, we found that strains expressing L5-tdTomato display the greatest fluorescence. Here, we describe detailed protocols for tdTomato-labeled M. bovis BCG imaging in real time for subcutaneous and pulmonary infections in living mice. These procedures allow rapid and accurate determination of bacterial numbers in live mice.
