Precipitation of 10X TBE - (Jan/19/2006 )
Hello,
Thanks for all the replies. This used to happen with 10X TBE we bought from Invitrogen. In addition some has left the labs in the summer and one of the items remaining was a 10L box of 10X TBE that was purchased from Sigma. The spigot was already inserted and when I tried to remove some yesterday there was a lot of white precipitate at the bottom. This was not through preparation as it was already bought prepared. My thoughts were it occurred after being opened and then left sitting around for months although I was unsure how to remove the ppt.
Well, TBE can be prepared as a 5x or 10x stock buffer, but the 10x stock will precipitate during storage.
I'm not sure why, but I have this feeling that stuff precipitates easier if the bottle is opened at least once. That is, say I made a 10x TBE stock and autoclave it. If I don't open the bottle, the stuff will not precipitate. But if I had opened the bottle, the stuff will tend to precipitate more often.
I've noticed the same thing with 3% glutamine too. Bottles not opened don't have any precipitate at the bottom. But as I open/close the bottles every time I prepare culture medium, the glutamine precipitates, even though storage conditions were otherwise the same (4 deg C). Hmm...
Just re-autoclave it when this happens, it'll be fine
I remember being told that precipitation wa due to bacterial contamination. And Autoclaved TAE or TBE never precipitates until it is opened. As well I am using 10X buffer from invitrogen and it starts precipitating only after opening. So If you fell really concerned about this precipitation autoclave your stock buffer and use it fast (or keep it sterile) after opening
hmm...I have a theory
I think usually you can see bacterial contamination; 'floaters' that aren't quite like precipitated salts, once they are around in any sort of numbers you see a few floaters and then shortly afterwards there is turbidity
I suspect it is a function of pH
someone early on said that perhaps it is EDTA coming out of solution, and this makes quite a bit of sense
whether or not it is the EDTA, or some other borate salt, I suspect pH is your culprit. pH would change every time you open the bottle too, and would not be dependent on whether or not there were critters growing. something like 10X TBE would not be overly susceptible to contamination; there are concentrated salts in there; but I bet a small change in pH could alter the balance between the salts and cause a precipitate. has anyone tested the pH periodically to see how it changes over time?
anyways, just a thought
I think usually you can see bacterial contamination; 'floaters' that aren't quite like precipitated salts, once they are around in any sort of numbers you see a few floaters and then shortly afterwards there is turbidity
I suspect it is a function of pH
someone early on said that perhaps it is EDTA coming out of solution, and this makes quite a bit of sense
whether or not it is the EDTA, or some other borate salt, I suspect pH is your culprit. pH would change every time you open the bottle too, and would not be dependent on whether or not there were critters growing. something like 10X TBE would not be overly susceptible to contamination; there are concentrated salts in there; but I bet a small change in pH could alter the balance between the salts and cause a precipitate. has anyone tested the pH periodically to see how it changes over time?
anyways, just a thought
Thanks once again for everyone's help. I have to agree with amikins and that it is not a bacterial contamination. Looks nothing like bacterial contamination and is definitely of a crystalline nature. I think once the bottle is opened the TBE is reacting with the air which subsequently is affecting the pH and causing one of the chemicals to come out of solution.
i think what amikins said is more likely as well, its not very common for bacteria to survive in 10X tbe... or SDS for that matter....
its more likely to be a pH problem....
It could be due to pH, but why would subsequent autoclaving re-dissolve the precipitate?
maybe because you heat much more than when you only put it in a water-bath to dissolve crystals.
then you might better get rid of all little crystals.
precipitation occurs when you have a crystal, which is difficult to obtain in a first step, but then when it's here, precipitation goes fast.