Tris base or HCl? - which one to use? (Jun/21/2010 )
I wish to prepare SDS-PAGE seperating gel. In my protocol, it is stated "1.5M Tris, pH 8.8". But there are Tris-HCl and Tris-base in my lab. May I ask which one to be used? Is there any special reason for the presence of this two types of Tris? To prepare Tris buffer at specific pH, can I just simply choose either type of Tris, and adjust with HCl/NaOH until my desired pH?
Thank you very much!
you can use either and use hcl or naoh to adjust the pH.
you can prepare equimolar solutions of both and mix them until you get the desired pH. this way concentration of the buffering species (tris) remains constant during adjustment.
If you start with Tris-HCl and neutralize with NaOH, you end up with excess salt in your buffer. I know you are aware of this; just pointing it out to the gallery. Your method of mixing amounts of equimolar Tris base and Tris-HCl to reach the desired pH is certainly my favorite approach.
Thank you for the response!
Just curious, if I start with Tris-base and adjust with HCl, is there any effect with the buffer I made?
Emm...both mdfenko and phage434 mentioned about mixing equalmolar of Tris-base and Tris-HCl to get desired pH. But I do not really understand this. May I ask is there any changes in Tris concentration if I:
1) make 1M Tris-base and adjust to pH 6.8 with HCl?
2) prepare 1M of Tris-base and 1M Tris-HCl, and then mixing them to reach pH 6.8?
I think both ways will end up with final 1M Tris, right?
No, your first method will dilute the solution with HCl. With concentrated HCl this may be a very minor issue, but it is difficult to control the titration with concentrated HCl, so the dilution can be substantial. You would have to start with higher than 1 M Tris, then neutralize, then adjust the final volume to achieve 1 M concentration. You don't need to do this final adjustment with the mixing approach, and you never have to worry about going "too far" with the titration.
Thank you for the answers. They really helped me alot