450 ml RPMI 1640 Biochrom T121
50 ml Fetal Bovine Serum (heat inactivated has benefit)
5 ml N-Acetyl
L-Alanyl L-Glutamine Biochrom K0202
5 ml Na Pyruvate (10mM – 1.10mg/ml)
0.5 ml Gentamycin (50 mg/ml) Biochrom A271-23
additional supplement (own experience):
5 ml MEM Vitamines (100x) Biochrom K0373
10 ml Amino Acids (50x) Biochrom K0363
Cells grow as a suspension culture and double approximately every 20-24 hours. Recommended cell density between 0.2 x 106 and 1 x 106 cells/ml. A recommended maintenance schedule is to split the cells 1:4 every other day.
Cells died rapidly in the original medium without Gentamycin. Heat inactivated Serum, Gentamycin and other supplements to the RPMI medium helped.
Cells grow very well in serum free medium Hybridomed (Biochrom F8055).