6 Fluorescence microscopes
Although it is not used routinely in histology laboratories, fluorescence microscopes are used extensively in research. They are particularly valuable for viewing live cells that have been labelled with fluorescent markers. For example it is possible to transfect cells with fluorescent versions of their endogenous proteins so that the cellular location of a particular protein can be identified. They are also used in a technique analagous to immunohistochemistry, called immunofluorescence, in which cells are stained with antibodies that are either directly coupled to fluorescent tags, or the primary antibodies are detected with a fluorescent secondary antibody. In all cases the cells or tissues are illuminated with a high intensity light of a suitable wavelength to excite the fluorescent tag. The emitted light is collected by the objective and viewed through the eyepiece as normal, but usually additional filters are placed in the path of the emitted light so that only light of the correct wavelength, forming the image is observed. The usual arrangement of a fluorescence microscope is using incident illumination, generating a dark field image (Figure 11).