Based on what they knew about the fertilization envelope (a protective layer that forms around the egg when a sperm fuses with it), steinhardt, shatten, and their colleagues hypothesized that changes in the distribution of ca2+ ions in sea urchin eggs are involved in the formation of the fertilization envelope. in sea urchin eggs (as in most eukaryotic cells) the concentration of ca2+ ions is much higher in the endoplasmic reticulum (er) than in the cytoplasm. to see how cytoplasmic ca2+ concentration changes in the egg during fertilization, the researchers injected a ca2+-specific fluorescent dye into the cytoplasm of unfertilized eggs. after adding sperm to the eggs, they observed the eggs with a fluorescence microscope. the following images show the changes in fluorescence that occurred after a single sperm entered the egg. the fluorescence of the dye increased with increasing cytoplasmic ca2+ concentration. the green color indicates the region of the cell with the highest fluorescence at that point in time.