Developing a Titration Kill Curve (G418, Hygromycin B and Puromycin)
Split non-transduced, confluent cells 1:5 in 10 ml DMEM + 10% FBS media.
Transfer 0.5 mL cell suspension into 24-well plate containing 500 µl of (media + drug).
- G418 Sulfate
- Hygromycin B
- Puromycin Dihydrochloride. 
Examine viability every 2 days.
Culture for 14 days.
Replace the media containing antibiotic every 3 days.
Use the lowest concentration of your antibiotic that begins to give massive cell death in 3 days and kills all the cells within two weeks.
A general starting point is usually 400 mg/ml G418 for HeLa cells and 200 mg/ml hygromycin for CHO cells.
In mammalian cells the optimal level of puromycin is typically around 1 mg/ml.
HeLa cells are often selectable with 500 mg/ml G418, 500 mg/ml hygromycin, or 2.5 mg/ml puromycin, and SHSY-5Y cells are often selectable with 600 mg/ml G418 or 200mg/ml hygromycin.
