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	...	This lab will take two weeks to accomplish. In the first week, we will collect and fertilize sea urchin gametes. One half of the embryos produced will then be subjected to a NiCl2solution until Friday morning. On Saturday, the embryos will be fixed in preparation for staining in week two. During lab next week we will stain the labs with Ig8 antibody, an immunofluorescent stain. We will use this stain to identify PCMs and developing spicules. ** We will be usingHytechinus variegatusfrom the Florida coast. The gametes and embryos can be left at room temperature in order to develop** Experimental Protocol A. Fertilization of Urchins See standard protocol. B. NiCl2Treatment 1)After fertilization has occured, transfer half of the embryos into a solution of 10mM NiCl2. Transfer the remaining half, the control, into a solution of ASW. 2)The two groups should be left at least 24 hours to develop. Track the development through the mesenchyme blastula stage. 3)The groups should be prepared for stain when the control group (ASW) reaches gastrulation and has formed the pluteus larva. C. Staining See standard protocol.