How to make gradient gel (10-20%), how to control the speed of flow? - (Mar/21/2013 )

I wanna to make 10-20% tris-tricine gradient gel myself. But how to control the speed of flow to make perfect gradient gel? Thanks.

-savetld-

For a standard gradient gel former: Flow rate out of the mixing chamber will determine flow rate between the two chambers (low % and high %). Just make sure that maximal possible flow rate between the two chambers is greater than the maximal possible flow rate out of the mixing chamber. Start with equal volumes and mix in the mixing chamber with a stir bar.

-doxorubicin-

we use the flow controller (flowcock? see the image) from intravenous apparatus (without the drip bubble). some others use a peristaltic pump.

just make sure that the gel doesn't polymerize before the pour is completed and that the pour speed does not overrun the gradient mixing.

-mdfenko-

mdfenko on Fri Mar 22 12:43:49 2013 said:

Thanks. But what u mean "pour speed does not overrun the gradient mixing". Last time I do the gradient gel just directly connecting the mixer and the casting chamber. This time I plan to try peristaltic pump, but which speed I could use and how to control the strength of string bar, I mean usually I just use very gentle magnetic strength to stir the bar in the mixer, or I should use stronger?

-savetld-

if you pour too fast then the solutions won't mix completely. the stirrer should be fast enough to ensure thorough mixing without aerating and/or foaming the solution. you may have to adjust the stirring speed as the gel pours.

i like to pour the gel at a speed where it finishes pouring in about 20 minutes. that way i don't overrun the mixing and the gel won't polymerize in the gradient apparatus.

-mdfenko-