i was following the protocol on the website that fulvio also found. it looks like if i just pour the powders into 800 mL of milli q water the solution becomes cloudy and the powders precipate or something. if u dissolve calcium chloride in milli q water and magnesium chloride in milliq water separately then add to the stock solution with the other reagents it stays dissolved.

yes its necessary to check the pH of the PBS.

yes, it is necessary to adjust the pH, and also keep in mind that you could end up with Ca-phosphate precipitates...wasting your PBS...why not using other buffers?PBS is not good with Ca-Mg

Hi Melissa, yes I also adjust the pH, but I also find it easier to make up my PBS from tablets. I find this helps with reproducible results. ref P4417-50TAB Sigma-Aldrich.

Does anyone know how long it takes for the reagents to dissolve when making PBS?

Yes, you must adjust the pH if you are preparing this buffer yourself or using, for instance, commercial liquid/tablets 10x preparations. PBS is really easy to prepare and dissolve, if you find any problem to dissolve all the components, probably you are doing something really wrong. Anyway, there are commercial PBS 1x Ca/Mg-free preparations ready to use you could use

from CSH protocols....I guess you'll found all the answer there:

http://cshprotocols.cshlp.org/content/2006/1/pdb.rec8247.full?text_only=true

i was following the protocol on the website that fulvio also found. it looks like if i just pour the powders into 800 mL of milli q water the solution becomes cloudy and the powders precipate or something. if u dissolve calcium chloride in milli q water and magnesium chloride in milliq water separately then add to the stock solution with the other reagents it stays dissolved.

yes Melissa is what I wrote before, sorry to not have make myself clear...so it goes: if you mix phosphate (K or Na) with CaCl directly it will form Ca-Phosphate that precipates (the cloudy solution that you see) and it is quite difficult to dissolve. While if you make a stock with Ca and Mg and subsequently add this to the phosphate it should not precipitate..should because it can make precipitate depending on: Temperature, exact measurement of Ca and Phosp, residual Ca on the milliQ water...etc...

I think now it is solved....