Is the bubble of SDS a problem in SDS-PAGE running buffer? - (Jul/22/2010 )
Hi, may I know if this is a good way to prepare 1X Laemmli SDS buffer?
Here is how I prepare it:
I will add Tris base, Glycine, and SDS together into a beaker and the chemcials are stirred until completely dissolved. Afterwards, I will adjust it with MQ water to final volume (10L).
However, during the stirring, considerable bubbles of SDS were generated. After I run the 2D gel, some of the spots became blurred. I did not add the bubbles of SDS into the electrophoresis cell during the run. Is this reason that the buffer does not have enough SDS?
So may i know how do you prepare this running buffer? I heard from another lab that he dissolves Tris and Glycine first, then adjust with water to final volume, finally add SDS and stir slowly until use..it seems that less bubbles are formed, do you think that the bubbles would really make any difference?
Thank you in advance.
Hi,
May be you want to know abour SDS PAGE running buffer but not Lamellis buffer,
I usually prepare before our earlier stock finish,
So i am usually preparing 5X concentrate for 5 Ls. I weight all components evening (just before leaving lab) and put about 4 Lts water and keep in that form no shaking or stirring. Next morning u can check everthing will be dissolved and make up the final volume.
In case u want to prepare immediately u can dissolve SDS seperately in some hot water and put it into solution of Glycine and Tris Base .
With Regards,
Mole .
Hi, Mole
Thx for the suggestions. I shall try next time.
Good luck.
mole on Jul 23 2010, 11:22 PM said:
May be you want to know abour SDS PAGE running buffer but not Lamellis buffer,
I usually prepare before our earlier stock finish,
So i am usually preparing 5X concentrate for 5 Ls. I weight all components evening (just before leaving lab) and put about 4 Lts water and keep in that form no shaking or stirring. Next morning u can check everthing will be dissolved and make up the final volume.
In case u want to prepare immediately u can dissolve SDS seperately in some hot water and put it into solution of Glycine and Tris Base .
With Regards,
Mole .
barnacleman on Fri Jul 23 02:48:50 2010 said:
Hi, may I know if this is a good way to prepare 1X Laemmli SDS buffer?
Here is how I prepare it:
I will add Tris base, Glycine, and SDS together into a beaker and the chemcials are stirred until completely dissolved. Afterwards, I will adjust it with MQ water to final volume (10L).
However, during the stirring, considerable bubbles of SDS were generated. After I run the 2D gel, some of the spots became blurred. I did not add the bubbles of SDS into the electrophoresis cell during the run. Is this reason that the buffer does not have enough SDS?
So may i know how do you prepare this running buffer? I heard from another lab that he dissolves Tris and Glycine first, then adjust with water to final volume, finally add SDS and stir slowly until use..it seems that less bubbles are formed, do you think that the bubbles would really make any difference?
Thank you in advance.
I make my running buffer right before I use it and there is a lot of bubbles there. Never cared about it though, as they are above the wells or on the outside of the cassette anyways.