2D electrophoresis - (Sep/22/2005 )

Hi !
I wanted to have some help about 2D electrophoresis! I would like to know how much protein I have to produce to do a 2D gel and to visualize proteins by Coomassie! (I use in vitro expression of protein)

Thanks!!!

Hello,

The answer to that question depends on the size of the gel you wish to run. If you're using a small size gel strip (compatible w/ Bio Rad Mini Protean gels or small size IPG strip/NuPage gel from Amersham), then you should probably be loading 50µg up to 500µg, though I've seen people get away w/ as low as 10µg (not recommended). If you're going for a larger gel, then 500µg up to 2mg is possible, though I suggest you visit the manufacturer's website for a more-specific answer.

Another thing to consider is the type of Coomassie staining you wish to use. If you use the conventional Coomassie R250 staining, you'll need to use more protein to see-low abundance proteins than you would if you used a colloidal coomassie stain, or a silver stain, or sypro ruby for that matter.

I hope this helps.

Jon
jonmike.reed@gmail.com