PhD student leaving for vacation has asked an undergraduate student to perform daily media changes for his iPSCs while he is gone. The culture is happening in 12-well plates where a volume of 2 mL is optimal. On Saturday, when changing the media, the undergraduate decides to add 4 ml of media to the dishes (instead of 2 ml) because he wants to skip lab and watch the Super Bowl on Sunday. He decides to add twice the volume of media (4 mL) to tide the cells over till Monday. However, when the graduate student returns on Monday, he finds that some of his cells have died. Your job is to determine whether the cells died due to a lack of oxygen. For the calculations that follow, diffusion and reaction occurs only in one direction. Also, assume that reaction only occurs at the
A PhD student leaving for vacation has asked an undergraduate student to perform daily media changes for
his iPSCs while he is gone. The culture is happening in 12-well plates where a volume of 2 mL is optimal. On
Saturday, when changing the media, the undergraduate decides to add 4 ml of media to the dishes (instead
of 2 ml) because he wants to skip lab and watch the Super Bowl on Sunday. He decides to add twice the
volume of media (4 mL) to tide the cells over till Monday. However, when the graduate student returns on
Monday, he finds that some of his cells have died. Your job is to determine whether the cells died due to a
lack of oxygen. For the calculations that follow, diffusion and reaction occurs only in one direction. Also,
assume that reaction only occurs at the cell-media interface. Use Michaelis-Menten type kinetics for oxygen
uptake. You may use the following information:
Ps = 150 mmHg (ambient oxygen tension)
K = 1.19 nmol / mL / mmHg (solubility of oxygen in medium)
D = 2 ×10−5cm2/s (diffusivity of oxygen through medium at 37 °C)
Km = 0.5 mmHg (partial pressure at which O2uptake rate is half maximal)
Vmax = 3 ×10−5nmol / s / cell (maximum oxygen uptake rate)
Cell viability is compromised at an oxygen tension of Pδ = 0.5 mmHg or below. The flux of oxygen due to
diffusion through the medium is given by
D*(K(Ps −Pδ))/δ
where δ is the height of the medium.
If the cells were seeded in 12-well plates (3.5 cm2) at 10,000 cells/cm2, then what is the critical volume
of media at which cells begin to die due to oxygen starvation? Was the volume of media added by the
undergraduate the cause of cell death?
Cell culture or tissue culture is the process by which cells are grown in a controlled environment, often away from their natural habitat.
The term "tissue culture" was coined by American doctor Montrose Thomas Burrows, and another term for this technique is micropropagation. Cells of interest can be extracted from living tissue and then maintained under carefully regulated conditions without the need for an incubator set at body temperature (37 °C).
These conditions vary depending on the cell type but typically involve using a suitable vessel containing a nutrient-rich medium or substrate that supplies essential nutrients such as amino acids, carbohydrates, vitamins, and minerals, as well as growth factors, hormones, and gases like CO2 and O2.
The physiochemical environment, including factors like pH, osmotic pressure, and temperature, is also controlled during the process.
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I am confused where the 0.002 came from, and how I could calculate the max volume media.
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