Fig. 2 | Communications Biology

Fig. 2

From: Evidence-based guidelines for controlling pH in mammalian live-cell culture systems

Fig. 2

The control and stability of pH in CO2/HCO3-buffered medium. a Effect of increasing [lactic acid] or [lactate] on equilibrium pH of medium (DMEM D5030) with 22 mM NaHCO3 (10% foetal bovine serum (FBS), 1% penicillin–streptomycin (PS)) placed in 5% CO2. b Effect of metabolic lactic acid production by DLD1 cells (seeding density 4,000 cells per well, growth area 0.32 cm2 per well) on the pH of medium (DMEM D5030; 10% FBS,  1% PS) containing 0–25 mM glucose (osmotically compensated with NaCl). Error bars omitted for clarity. c Effect of varying starting glucose concentration on net glucose uptake and lactate production probed on the seventh day of incubation. 90% of glucose is metabolized to lactate. d Relationship between lactate production (measured by biochemical assay) and total acid production (calculated from the pH change and buffering capacity). Slope of 1.0 indicates that medium acidification is due to lactic acid production. e Cell growth of three colorectal cancer cell lines (seeding density 4,000 cells per well, growth area 0.32 cm2 per well) measured from protein biomass (sulforhodamine B (SRB) assay) after 6 days of incubation in DMEM (D7777; 10% FBS, 1% PS, 25 mM glucose) over a range of starting pH attained by varying [HCO3] at constant 5% CO2. Data are normalized to the optimum pH derived by best fit to biphasic curve. Optimal growth is near the physiological pH of 7.4. f Effect of varying pCO2 on medium pH, mimicking the withdrawal of medium from under CO2 incubation. All experiments were repeated three times (three technical replicates each). Data are shown as mean ± SEM

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