Proper ratio of stacking and separating gel - (Apr/25/2008 )
Hello everybody,
it could sound a little bit strange. My problem is that I have a large volume of a protein sample. But my capacity of the gel pockets are restricted. Now my question can I pour a gel with a long stacking gel that my volume will fit properly? Because I don`t want to precipitate my samples. Or will I get problems? I am not sure. Once I read one should have a long separating gel. My mw of my protein is about 180kDa.
Please help me.
Thanks
Tiffy
it could sound a little bit strange. My problem is that I have a large volume of a protein sample. But my capacity of the gel pockets are restricted. Now my question can I pour a gel with a long stacking gel that my volume will fit properly? Because I don`t want to precipitate my samples. Or will I get problems? I am not sure. Once I read one should have a long separating gel. My mw of my protein is about 180kDa.
Please help me.
Thanks
Tiffy
Yeah of course, the length of the separating gel should be greater than the the stacking gel. Instead of increasing the length of the stacking gel, it would be better if u concentrate ur protein sample by dialysis, so that the volume of the sample to be loaded in the gel will become minimum.
depending on how much more volume you need, you might try cutting your comb to get larger wells, potentially increasing the size by about 50% or so.
Seemed you are trying to get a bigger gel pocket to fill your sample. When you are pouring the gel by yourself, use a tape to wrap 2 or 3 wells together on your comb. This will make a well 2-3 folder bigger than the regular one. Of course it will give less wells for your samples. Hope it helps.
Instead of precipitate the sample, you can also try concentrate it by columns.
it could sound a little bit strange. My problem is that I have a large volume of a protein sample. But my capacity of the gel pockets are restricted. Now my question can I pour a gel with a long stacking gel that my volume will fit properly? Because I don`t want to precipitate my samples. Or will I get problems? I am not sure. Once I read one should have a long separating gel. My mw of my protein is about 180kDa.
Please help me.
Thanks
Tiffy