Mongelard Fabien wrote:
>> How do you prepare 2 M sodium acetate, pH 4 used in RNA extraction ?
> Do you dissolve 2 moles of Na Acetate and bring the pH to 4 with acetic
> acid (as indicated in Maniatis to prepare 3M NaAcetate pH 5.3 for
> exemple), or do you start with acetic acid, and adjust the pH using NaOH ?
When making up buffers like this I usually refer to Data for Biochemical
Research by Dawson, Elliot, Elliot & Jones (Oxford Science
Publications). They have a series of recipies for buffers based on
adding the correct ratios of acid and base according to the Henderson
Hasselbach equation. To make up 2M Na Acetate pH 4.0, first make up 2M
NaAcetate 272.2 g/l and 2M Acetic acid (calculate the molarity based on
the density if using glacial) it should work out as 115 ml of glacial
added to 885 ml water. Then add 18.0 ml 2M NaOac to 82.0 ml 2M acetic
acid. This should give you a solution which is 2M with respect to
acetate and is pH 4.0. Make sure you check the pH after making it up in
case I've mis-calculated or made a typo!
> Results are different with respect to Na+ and Acetate concentration.
You are trying to make a buffer based on the acetate <=> acetic acid
equilibrium therefore it is the concentration of these species that you
need to worry about and that need to total 2M. The Na+ is just a
I hope this helps,
The Sanger Centre
email: rab at sanger.ac.uk Wellcome Trust Genome Campus
phone: (01223) 494949 Hinxton
fax: (01223) 494919 Cambs