Hepes buffer solution

Phoenix^TM was pre-heated to 37°C at least 30 min prior to each experiment. The cell container was sterilized with 70% isopropyl alcohol then stored in an open position inside a standard incubator enriched with 5% CO₂. Prior to loading, media within t-flasks for both incubator treatment groups were supplemented with 25 mM of HEPES buffer solution (VWR) to buffer solution pH without the use of CO₂. Once seeded t-flasks were added, the cell container lid remained in an open position. After 30 min of incubation, the cell container lid was moved to a closed position while inside the standard incubator. The cell container was then transferred to Phoenix^TM. Ground transportation occurred between Kahului and Haiku, Hawaii, United States, where Phoenix was loaded inside a motor vehicle then transported 13 miles twice per day. Air transportation occurred in Kahului, Hawaii, United States. Phoenix was loaded into a motor vehicle and driven 4.2 miles to Maui Flight Academy at the Kahului International Airport. After loading Phoenix^TM into the cargo hold of a Cirrus SR22, a ∼30-min test flight was performed that included two take-offs (2.1 g), two 45° bank turns (1.6 g), two 60° bank turns (2.0 g), nosedive (0 g), and two landings (0–2.5 g). Additional values can be found in Figure 8. All flying aerobatics were performed at an altitude of 1500 ft.

MDA-MB-231 cells were obtained from LGC promochem (London, UK) and maintained in high glucose Dulbecco’s Modified Eagle’s medium (DMEM, Gibco, 31966–021) supplemented with 20 mM HEPES buffer solution (1 M, VWR Life Science, J848-500 mL), 1 × MEM Non-Essential Amino Acids (MEM-NEAA, Gibco, 11140–035), 1 × Penicillin/Streptomycin solution (Gibco, 15140–122) and 10% heat-inactivated Foetal Bovine Serum (FBS, Gibco, 10500–064), at 37 °C and 5% CO₂. Cells were routinely passaged at 90% confluence by washing with Dulbecco’s phosphate-buffered saline (DPBS) and harvesting with 0.05% Trypsin–EDTA (both Gibco, 14190–144 and 25300–104, respectively).