We describe labeling of bacteria with amino-specific or sulfhydryl-specific Alexa Fluor dyes, methods that allow visualization of flagellar filaments, even in swimming cells. Bacterial flagellar filaments are long (~10Â μm), but of small diameter (~20Â nm), and their rotation rates are high (>100Â Hz), so visualization is difficult. Dark-field microscopy works well with isolated filaments, but visualization in situ is hampered by light scattered from cell bodies, which obscures short filaments or the proximal ends of long filaments. Differential interference contrast microscopy also works, but is technically difficult and suffers from a narrow depth of field and low image contrast; background subtraction and contrast enhancement are necessary. If filaments are fluorescent, they can be imaged in their entirety using standard fluorescence microscopes. For imaging in vivo, blurring can be prevented by strobing the light source or by using a camera with a fast shutter. The former method is preferred, since it minimizes bleaching.
CITATION STYLE
Turner, L., & Berg, H. C. (2018). Labeling bacterial flagella with fluorescent dyes. In Methods in Molecular Biology (Vol. 1729, pp. 71–76). Humana Press Inc. https://doi.org/10.1007/978-1-4939-7577-8_7
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