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Fig. 2. Schematic overview of SMM applied to a living zebrafish embryo. A zebrafish embryo was injected with an expression vector for a YFP fusion protein shortly after fertilization. At 2 dpf, it was placed on a coverslip coated with poly-L-lysine in a drop of egg water. The tail region of the embryo was covered with a 0.75-mm-thick agarose (2%) sheet. On the lower left part of the figure, an epifluorescence picture is presented of the outer layer of the epidermis, showing the fluorescent signal of YFP-C10H-Ras in the cell membranes. Morphologically, the cells in this layer are homogenous and are characterized by a hexagonal shape. On the lower right part, a TIRFM image of an area within the field of view of the epifluorescence picture is presented, with examples of individual YFP molecules shown in circles. Scale bars: 5 µm.