Pen strep nystatin

Cell lines used: Expi293 (ThermoFisher Scientific), TK-1 (ATCC), HEK293 (ATCC). All cell lines in the laboratory were tested every 2 months for mycoplasma and discarded when positive. Expi293 cells were grown in Expi293 Expression Medium (ThermoFisher Scientific, USA) in disposable Erlenmeyer flasks at 37°C and 8% CO₂ on a shaker rotating at 125 rpm. Cells were grown at densities between 0.3x10^{6 (link)} – 5x10^{6 (link)} cells per mL. At least 3 passages after thawing, cells were transfected with the expression plasmid encoding the MAdCAM-D1D2-Fc using the Expifectamine293 transfection kit (ThermoFisher Scientific, USA). 18 h post transfection, enhancer 1 and enhancer 2 from the Expifectamine293 transfection kit were added to boost recombinant protein expression levels. Five days post transfection the culture medium was harvested for purification of the secreted protein.
TK-1 cells were grown in RPMI1640 medium supplemented with 10% fetal bovine serum, L-glutamine and Pen-Strep-Nystatin (Biological Industries, Israel). Cells were grown at densities between 0.3x10^{6 (link)} – 2x10^{6 (link)} cells per mL in either T25 or T75 cell culture flasks (Greiner bio-one, Austria).