5.1 Fluorescence microscopy allows observing specific structures individually and combined

Fluorescence microscopy makes it possible to observe specific cellular structures both individually and in combination. After staining different structures within a cell using fluorophores of different colours, the microscope’s separate channels can be used to select specific wavelengths of light to excite each dye one at a time. This allows for the capture of individual images, each showing a single labelled structure (Figure 14a and b, and Video 1). These individual fluorescence images are then digitally merged or overlaid using computer software, creating a combined image that reveals the spatial relationship between all stained structures within the cell or tissue (Figure 14c).

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Figure 14 (repeat of Figure 11) HeLa cells labelled with two fluorescent dyes: one that binds to DNA and one that accumulates inside mitochondria. (a) Micrograph taken after illuminating the cells with ultraviolet (UV) light to excite the dye bound to DNA, which then emits blue light. (b) A second micrograph taken after illuminating the cells with green light to excite the dye localised in mitochondria, which then emits red light. (c) The images are merged using computer software to show the fluorescence of both dyes at the same time.

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The figure consists of three parts, Figures 14a to c, that illustrate micrographs of HeLa cells labelled with two fluorescent dyes.

Figure 14a: a fluorescence micrograph shows an oval-shaped nucleus emitting blue fluorescence. The scale bar is 10 micrometres.

Figure 14b: a fluorescence micrograph shows a cluster of mitochondria arranged in a circular pattern emitting red fluorescence.

Figure 14c: a merged image from parts a and b shows an oval-shaped nucleus emitting blue fluorescence and surrounded by mitochondria emitting red fluorescence.

Figure 14 (repeat of Figure 11) HeLa cells labelled with two fluorescent dyes: one that binds to DNA and one that accumulates inside ...

Capturing separate images and merging them allows detailed studies of the localisation of structures and proteins. You will see more examples when using the digital fluorescence microscope yourself.