I need to culture mammalian cells at low calcium 0.03mM and high calcium 1.8mM condition. There is only one suitable calcium-free medium available in the market (MEM, low glucose, no hepes). Then I add 10% FBS, vitamins, essential amino acids, glutamax and calcium chloride to achieve similar formula as DMEM. When I add the calcium chloride first to the MEM, the high calcium medium turns turbid after some hours and I saw white precipitate at the bottom after a few days. If I add FBS first, there is no precipitate. But when I incubate this medium in 37oC 5% CO2, it turns turbid after one day and precipitates are found after some days, even without cells inside. There is no turbidity or precipitate at all in the low calcium medium, so definitely this is related to calcium chloride concentration. I measured the pH of media, before incubation both are pH 8.0, after incubation both are around pH 7.1-7.2. My DMEM (low glucose, no hepes) has a 1.8mM calcium concentration and pH 8.0 before incubation and pH 7.4 after incubation. My cells could still grow very fast in the turbid high calcium MEM but after a few passages they slow down a lot and the morphology looks very different.
Hello. thank you very much for your informative reply. Finally I found out the reason of the precipitates. It is because of the high phosphate concentration of MEM. It is 10 times more than DMEM!!!! so the calcium phosphate precipitated out. I checked the maximum concentration of calcium ions I can have without precipitates is only 0.3mM. so I cannot achieve high calcium condition with this medium. Now I have to change my experimental design.