Isolation of mouse embryos can be used for: (1) Culturing mouse embryos in vitro and figuring out appropriate culture conditions. (2) Using embryos for developmental biology research.
Operation method
Separation of mouse embryos
Principle
An embryo is a young body within the first two months of pregnancy. Blastocyst, early embryo development and zygote development are a continuous process. During the somite period, all three germ layers undergo changes. The ectoderm is depressed in the midline of the back into a groove called the neural groove, and the banks of the groove are called the neural ridges. The neural ridges gradually approach and heal, causing the nerves to evolve into a longitudinal neural tube running the length of the embryo. The neural tube and the neural ridge evolve into the nervous system in the future. The endoderm curls with the embryo to form a tube, called the proto-gut, which is divided into the foregut, midgut and hindgut according to its location.
Materials and Instruments
DBSS Move 1. Mating. Mating is carried out by combining males and females in separate cages, with females entering estrus after 3 days, when mating is most successful. This process can be planned so that embryos are produced at the right time. In order to determine the time of successful mating, the female's pessary should be examined every morning. Caveat 1. When instruments are immersed in ethanol and then ignited, i.e., sterilized, great care should be taken not to put the still hot instruments back into the ethanol!2. All of the above steps should be performed outside of the tissue culture room, a quick procedure in a small ultra-clean bench will help maintain sterility. Do not bring live animals into the culture room as they can introduce contamination. If animal carcasses are to be disposed of in the culture room, they must be immersed in or coated with ethanol and disposed of as soon as possible after use. Common Problems The development of the embryo is an extremely delicate and complex process of differentiation of cells and tissues in a certain order, and disturbance of any part of the process can lead to various malformations. Especially when organs are rapidly differentiating and developing, they are most vulnerable to the interference of teratogenic factors. For more product details, please visit Aladdin Scientific website.
70% ethanol
Petri dish Sharp tweezers Sharp scissors Ultra-clean bench Bunsen burner
2. Calculate the time of embryo development: The day when the pessary is found is designated as day zero, and the time of embryo development is calculated from this day onwards. The entire gestation period is about 19~21 days. The appropriate time to isolate the complete embryo for cell culture is the 13th day, when the embryo is already quite large but still contains a large number of undifferentiated mesenchymal stromal cells, which are the main source of cultured cells. However, a license may be required (e.g., in the United Kingdom) to isolate and process embryos more than halfway through gestation, so it may be preferable to use embryos that are 9 to 10 days old. Although the number of tissue cells isolated from such embryos is relatively small, the proportion of cells capable of growth is high. Most organs except the brain and heart begin to form on day 9 of gestation and are difficult to isolate before day 11. Separation of single organs is easier on the 13th and 14th days, and most organs are fully formed by the 18th day. 
3. Mice were executed by neck-pulling (U.K. Schedule I procedure) and the abdominal surfaces were carefully disinfected with 70 % ethanol. 
4 . The skin is laterally torn at the midline of the abdomen near the diaphragm, and the skin is flipped on both sides to reveal the uncontacted abdominal wall. 
5. A longitudinal dissection along the midline was made with sterile scissors to expose the abdominal organs, and the embryo-filled uterus was visualized in the posterior abdominal cavity. 
6. The uterus is removed and placed in a 25 ml or 50 ml screw-top flask containing 10 ml or 20 ml of DBSS (DBSS for sampling (50 ml of BSS containing a high concentration of antibiotics in a sterile beaker used to cool burned instruments) in 25-50 ml screw-top test tubes or a general-purpose container). 
7. Transfer the untreated uterus into the culture chamber to a new petri dish containing sterile DBSS. 
8. Separate the embryos:
(a) Tear open the uterus with two sterile forceps, taking care to keep the tips of the forceps closed to avoid excessive disruption of the uterus and excessive pressure on the embryo. 
(b) Peel the embryo from the membranes and placenta and place it on the other side of the petri dish, avoiding blood. 
9. Transfer the embryos to a new Petri dish or, if a large number of embryos are to be removed (e.g., more than 4 or 5 clutches), place the Petri dish on ice. 
