Preparation of Primary Cultures of CEFs

1. One dozen eggs (appropriate strain), incubated for 10 d. Remember to turn the eggs 1/4 turn every 2 d to increase viability.

2. Sterilized surgical instruments (scissors, watchmaker's forceps, razor blade).

3. Sterilized 250-mL Erlenmeyer flask.

4. Sterile 10-mL wide-mouth pipets.

5. Sterile 50-mL plastic centrifuge tubes.

6. Sterile 1.5-mL cryovials.

7. 10-cm Petri and tissue-culture dishes.

9. 1X Trypsin/EDTA (0.5 mg/mL trypsin, 0.2 mg/mL EDTA), available as 1X stock in Modified Puck's Saline A from Gibco-BRL (Gaithersburg, MD).

10. Fetal calf serum (FCS), available from Gibco-BRL.

11. Chicken serum (CS), available from Gibco-BRL.

12. Penicillin/streptomycin (pen/strep), available from Gibco-BRL.

13. Dulbeco's Modified Eagle's Medium (DMEM; high-glucose, l-glutamine, sodium pyruvate), available from Gibco-BRL.

14. Dimethyl sulfoxide (DMSO), available from Sigma. DMSO is an irritant requiring adequate safety precautions.

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