does anyone use pre-chilled SDS buffer? - (Apr/26/2008 )
I use chilled transfer buffer and an ice pack for doing the transfer, but does anyone use pre-chilled SDS buffer for SDS-PAGE? If the gel gets hot it lowers the resolving power, making bands fuzzier...my bands are fine, but maybe they could be even sharper. Has anyone else tried this? I do my transfer at constant 120mA, and run the SDS-PAGE at constant 30mA for about 1 hour, maybe that is too low to need pre-chilled buffer.
you mean SDS running buffer? it is recommended to chill during running; some chambers use large volume of SDS running buffer or water to prevent warming during run; Hoefer´s Mighty Small chamber, f.i., can be cooled by running water; beware that at 4°C SDS tends to precipitate!
I believe in this issue it's the concern to prevent warm/hot instead of keep it cold/freezing. Honestly I never see bands become sharper when running in ice-cold condition. So it might be easy just using the running water to prevent the warming problem.
Yes , we use pre-chilled SDS running buffer....especially with the new biorad's electrophoretic system in which you can simultaneously run 4 gels at once...things do get heated up a lot. To be honest we always store it at 4 deg but never experienced a problem with SDS precipitation.
We started doing this to make a chemiluminiscent marker (containing peroxidase enzyme) work better. but as such never experienced problems with heated up gels/ buffer during electrophoresis..
I think it is more important to keep things cooled during transfer.
SDS running buffer will precipitate if store at 4degree for more than one month.