Part A - Experimental technique: Tracking cytoplasmic Ca2+ distribution after a sperm enters the egg
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.
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