Animal Cell Culture: A Guide to Basic Experiments

Chicken embryos are easier to isolate because they are larger than contemporaneous mouse embryos. As with mouse embryos, chicken embryos provide primary cultures of mesenchymal cells for cell proliferation analysis, are used as feeder layer cells, and can be used as a substrate for viral propagation. Because of its large size, the chicken embryo is also more likely to isolate single organs for the production of specialized types of cells, such as hepatocytes, cardiac muscle, and lung epithelial cells. The use of chicken embryos is also subject to animal regulations (e.g. in the UK) and a license is required for the use of embryos more than halfway through gestation.

Operation method

Scheme 12.2 Experiments in isolating chicken embryos

Principle

Chicken embryos were aseptically removed from the eggs and placed in petri dishes.

Move

Material:
Aseptic material
DBSS: BSS (BSS containing high concentrations of antibiotics, see Appendix I) in 25-50 ml screw-top tubes or general purpose containers for material extraction
BSS: 50 ml in a sterile beaker for cooling burned instruments.
20-50 ml small beakers or egg cups
Straight and curved forceps
9 cm Petri dish

non-sterile material
Fertilized eggs incubated for 10 days
70%ethanol
cotton swabs

Incubator (not C02)

procedure

1. Incubate the eggs in a humid environment at 38.5°C: rotate them 180° every day. Chicken eggs are also incubated for 20 to 21 days, but they are not at the same stage of development as mouse embryos. Isolation of the whole embryo for cell culture takes place at about day 8. If organs are isolated, this is done at about day 10 or 13 (day 10 is the deadline for a license in the UK).

2. Sterilize the eggs with a 70% ethanol swab and place them blunt end up in a small beaker (Fig. 12.4a).

3. Use sterile forceps to break the eggshell (Fig. 12.4b) and peel the eggshell to the edge of the air flag (Fig. 12.4c).

4. after re-eliminating the forceps (immersed in ethanol, dried in ethanol, and cooled in sterile BSS plants) peel off the white shell membrane with forceps, exposing the chorionic adventitia membrane (CAM) and blood vessels underneath (Fig. 12.4d, Fig. 12.4e).

5. Prick the chorionic villous membranes with curved forceps (Fig. 12.4f), gently clamp the underside of the head and lift the embryo (Fig. 12.4g, Fig. 12.4h), do not close the forceps completely to avoid the neck being cut off and place the pad of the middle finger underneath the forceps and use it to limit the pressure on the index finger (Fig. 12.4g).

6. place the chick embryo into a 9 cm diameter petri dish containing 20 ml DBSS (Fig. 12.4i) and wait for further dissection and culture (see Scheme 12.7).

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