Procedure for dissecting day 7 embryos for analysis in whole mount. The decidua are not removed from the uterus, allowing for more rapid dissection. Two pair of fine forceps (Dumont No. 5 or No. 5 Biologie forceps, Fine Science Tools, Foster City, CA), disposable petri dishes, a dissecting microscope and cold PBS are required.
Place the dissected uterus in small petri dish and cover with cold PBS. Arrange the uterus such that the membrane that attached the uterus to the body wall (the mesometrium) is facing away from you (A). Under a dissecting microscope grasp the uterus immediately next to the deciduum with both pairs of forceps and tear the uterus off the top of the deciduum--the uterus is tough and muscular, whereas the decidual tissue is soft and spongy (B). With one pair of forceps, clip off a portion of the exposed deciduum, amounting to about one quarter to one fifth of the entire deciduum (C and D). The tip of the embryo (midventral or distal tip) should now be exposed to view (D). Reichert's membrane is usually still attached to the embryo at this point. To make a hole in the nearly invisible Reichert’s membrane that covers the embryo, insert a pair of forceps immediately adjacent to the embryo, close and remove (E). Squeeze the embryo out of the deciduum and Reichert’s membrane by placing the forceps on the overlying uterine wall and applying gentle pressure (E). The embryo will usually pop out intact, without Reichert’s membrane and the ectoplacental cone. If they remain attached, they should be removed by careful dissection.
This method of dissection is rapid and is well suited for preparing embryos for whole mount in situ hybridization, immunocytochemistry, LacZ staining or TUNEL staining. It is not appropriate for whole embryo culture.
A Conlon Lab Protocol
pdf file for this protocol