11 replies to this topic

[Ecoli0157]

I have been having some trouble with contamination of my plates.
I swirled 10X PBS stock that I used to make inoculum and saw particles that form tornado-like shape.
Looks similar to bacterial growth, but I don't think bacteria can grow in it without any nutrient. Its just salt solution.
So, I was wondering it may be just the salt precipitating out of solution upon temperature fluctuations or something.
And if it is really the salt precipitation, can I just use it like that? Re-autoclave? Filter?

I await your suggestions.

Thanks

[BHARGAVI]

It has been observed by me that sometimes PBS preparation contains spores. But in your case, there is little possibility of growth of any spores as there is no nutrition. One thing you can do to avoid the presence of precipitates of salts or colony like tornado-like shapes, is you can filter the PBS just before use with 0.5 or 0.2 micron filter.

Heat sterilization sometimes cause the salts of PBS to precipitate out. Better avoid it.

[molgen]

Cyanobacteria and single cell algae can grow in the PBS.
I don't think that thy can in x10 because of the osmolarity.

We always filter sterilize it in any case.

[seashell83]

I have a question here, i recently made a 10x stock of PBS like i do usually and after making it i filter sterlized it using a 0.2 micron membrane and kept it at 4 degree in a clean container, the next morning i saw formation of crystals in my PBS which has never happened before - so now i tried to mix the crystals back into solution but its not goin in even being a RT for an hour... - HELP !!!!!!!!!!!!

[seashell83]

The PBS i made is without Ca or Mg

[BHARGAVI]

Its nothing but salt precipitates. The way you add and dissolve the salts and sequence of addition also matters for the quality of PBS.

[aimikins]

I've seen bacterial contamination in PBS.  it's true that there's not much in there, but you can still see contamination.  I would FS your stocks for sure, or autoclave.

[perneseblue]

seashell83, on Mar 26 2009, 08:03 AM, said:

I have a question here, i recently made a 10x stock of PBS like i do usually and after making it i filter sterlized it using a 0.2 micron membrane and kept it at 4 degree in a clean container, the next morning i saw formation of crystals in my PBS which has never happened before - so now i tried to mix the crystals back into solution but its not goin in even being a RT for an hour... - HELP !!!!!!!!!!!!

Try putting your 10x stock PBS into a water bath at 68C or a hybridisation incubator set at 68C. It should go into solution

[rkay447]

PBS is so cheap and easy to make, you might have gotten a decimal point off and added too much of one of the salts.  It's much easier, faster and safer (in the sense of your experiments working) to get rid of the questionable PBS and make a new batch.

[molgen]

seashell83, on Mar 26 2009, 06:03 PM, said:

I have a question here, i recently made a 10x stock of PBS like i do usually and after making it i filter sterlized it using a 0.2 micron membrane and kept it at 4 degree in a clean container, the next morning i saw formation of crystals in my PBS which has never happened before - so now i tried to mix the crystals back into solution but its not goin in even being a RT for an hour... - HELP !!!!!!!!!!!!

PBS is so cheap!!!
Why don't you just buy it??

[memo]

Be sure you use good water to dissolve the salts of 10x PBS.
After autoclave sterilization I once saw a precipitate! It seems that the DI water installation has broken down and my water contained to much carbonates or something!

At 4 degrees celsius your salts precipitate and form christals. Heat it up again and they dissapear.

[mdfenko]

are you sure that the glassware that you stored it in is not contaminated with cations or detergents?

if there is calcium present (from rinsing with tap water instead of deionized, or if the deionizer is exhausted) then you may be seeing calcium phosphate precipitating (or magnesium phosphate if it was heated).