2.1 Cells and media
CHO-K1 (RIKEN Cell Bank, Tsukuba, Japan) and recombinant CHO cells were
cultured in F12 medium (#N6760; Sigma-Aldrich, St Louis, MO, USA)
supplemented with 10% fetal calf serum (BioWest, Nuaillé, France) and
antibiotics (Penicillin-Streptomycin, #15140122; Invitrogen, Waltham,
MA, USA). Cells were cultured in a 5% CO2 incubator at
37°C. For serum-free cultures, CD CHO AGT Medium (#12490025,
Invitrogen) containing 4 mM L-glutamine (#074-00522; Fujifilm Wako Pure
Chemical Corporation, Osaka, Japan), 0.2% anti-clumping reagent
(#0010057AE, Invitrogen), and antibiotics (penicillin-streptomycin) was
used. When cells were cultured in serum-containing medium, 100-mm cell
culture dishes (BioLite #130182; Thermo Scientific, Waltham, MA, USA)
or 24-well tissue culture plates (BioLite #130186, Thermo Scientific)
were used. For serum-free suspension culture, bioreactor tubes (#87050;
TPP Techno Plastic Products AG, Trasadingen, Switzerland) placed in a
shaker (Model #0081704-000; Taitec, Koshigaya, Japan) with a 45° angle
were incubated at 37°C in a 5% CO2 incubator.