Fig. 2

a. Picture of a metaphase II mouse oocyte with the picture of a micrometre grid (10-μm intervals) superimposed. b. Zygotes were filmed during injection with a gentle flow of concentrated suspension (0.2 mg/mL), but only up to the filling of the perivitelline space with an injected volume ‘x’; selected frames were extracted from movie at indicated time points. Note that the perivitelline space is completely filled at 25 s, but recovers partly after 2 min and completely after 4 min. c. Serial doubling dilutions of a standard of green-fluorescent dextran beads (Oregon Green dextran beads, OGDB). Zygotes were blown up with a maximum flow of OGDB suspension applied continuously for 30 s, for each of six concentrations (0.2 mg/mL halved by serial dilutions down to 0.00625 mg/mL), causing the zona to be evacuated and the cytoplasm to be completely replaced by OGDB. The green fluorescence intensity was recorded for each dilution with the same excitation and time exposure (d). Dilution factor: A zygote injected as in (B), shown in the small inset photo, had a fluorescence intensity corresponding to 42% (1/2.4) of the fluorescence of the standard alone. These data allow one to solve a simple equation for the injected volume ‘x’: dilution factor = 2.4 = (x + 220 pl) / x = 157 pl. Size bar, 50 μm. AU, arbitrary units of fluorescence intensity