I've been trying to prepare a cell culture media (1% BSA) with palmitic acid (PA), but it keeps precipitating.
I successfully made a stock by dissolving 1 g PA in DMSO, so that's not a problem here. BUT when trying to add the stock into my medium (Both in 37 C), it gets solid! I'm adding 33 ul PA-DMSO in 50 ml medium, so it's not even a particularly high concentration I'm trying to make... I tried vortexing and kept it in +37 C for an hour, but it didn't show any signs of dissolving.
Next, I red some people heating up the medium and PA stock to 70 degrees before mixing, and while that helped (although there are still tiny flakes floating!), I don't think that's any good for the proteins of the medium (e.g. BSA). So, how is this supposed to be done? And is it expected that there will always be some tiny flakes no matter how prepared?
Hello Sanna Kettunen
You may try the following method.
You may make 200mM stock of Palmitic acid in ethanol.
You may dissolve 51.2mg Palmitic acid (Mol.wt. of Palmitic acid is 256.43 g/mol) in 1ml ethanol (100%) to give a stock of 200mM.
Then take 40ul of the above stock solution and add to 1.96ml of 10% fatty acid free BSA (in culture media) to give 4mM diluted stock of Palmitic acid. The solution must be shaken overnight at room temperature.
The following day, you may filter sterilize the diluted stock using 0.22μM filter. If required, further dilutions to be used in cell treatment may be made in culture media.
Best.
Malcolm Nobre Thanks for the suggestions! So do you think ethanol is the key here? Or shaking o/n? As mentioned, I’ve used DMSO and it works fine for the stock but the following step with media seems to be the problem. Haven’t tried overnight shaking tho!
Well, now I tried shaking overnight but it didn’t do anything; PA it’s still as visible (and big!) particles :/
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