BN-PAGE can be really tricky to master. I can recommend the excellent article by Ilka Wittig and co-authors I put into the attachment. It will give you a wealth of information.

I think it is highly recommended that you cool your gel during electrophoresis in order to preserve protein-protein interactions. The errors your power supplies gave you might be due to other problems with the electrophoresis chamber (internal buffer leaking from one tank to the other leading to a short-cut, etc.).

Good luck!

Thank you for the paper Christian. It is the protocol I am following. Seemed easy on paper, much more nuanced in real life :)

Hi Natasha,

I came across the same issue of the power supply not liking the cold-room. I think the issue is the high resistance in the BN system. My power supply throws an error if the current drops below 10 mA and that would happen within half an hour or so in the cold-room, but not at room temperature (although I don't really understand why!). If you have the money, there are specialized low current power supplies which should be ideal. Also, I think newer power supplies (the Bio-rad ones at least) have the option of turning off the no load detection so they'll keep running even if the current is below the cut-off threshold for the error message, so it's worth checking the manual for the particular power supply that you're using.

If neither of these is an option, the way I keep my gels cold and my power supply happy is to do it at room temperature, but put the gel tank into a big polystyrene box and pack it with ice.

As for seeing the samples, some protocols suggest to change the cathode buffer to one with a tenfold reduced (or zero) Coomassie concentration after an hour which will definitely help with seeing where your samples are! This is a hassle though, so the way I did it was to measure how long it took for my Coomassie front to reach the end of the gel the first time I did it, by pouring off the blue buffer and replacing every 15 minutes. Now I just set my gels to run for that amount of time (1hr 45mins for 8% 0.75 mm minigels at 180V).

Hope this is helpful!

Phil.

Thanks Philip. Very informative!

I solved the power supply issue by putting it in the cold room, turned on, the day before running the gel (as opposed to a couple of hours since all our power supplies are normally at RT). It was really unhappy at first (lots of not so good sounding noise) but then it quieted down and worked perfectly.

I realize that this is an older topic, but  my Power Supply (BioRad PowerPac 200) does indeed run into the same issue at room temperature, which is only happening when running BN-PAGE gels (constant voltage 150V as recommended for the system).

It always starts up fine again. I would recommend to keep an eye on your Power Supply for the final half of the run if you are having this problem.