working out percentages - (Mar/20/2006 )
I was wondering of anyone could help me. I was wondering how you work out the percentage of an sds-page gel - for example if a gel is a 10% gel, it is 10% of what in what? I would like to be able to work out things like how much of everything to add if I wanted to make a particular volume of a particular percentage (at the moment i have two recipes for a 10% gel, each with different final volumes - i want to be able to work out if they are comparable) - can anyone shed any light?!
let's assume that your stock of acrylamide is 30%
if you want to prepare a 10 % gel, you will have to dilute 3 times your acrylamide stock.
for 10 mL of gel, you will have to take 3.33 mL of acrylamide.
some stocks are 40%, that's why you may have different protocols, depending which initial concentration of acrylamide you have.
And you dilute your acrylamide in a solution containing Tris-HCl pH 8.8 0.375M, SDS 0.1% final concentration
Then to polymerize you have to add APS 0.05% and temed 0.05%
this is for the separation gel.
for the stacking gel I dilute in Tris-HCl pH6.8 0.25M, SDS 0.1%
to polymerize I add APS 0.05% and temed 0.1%
If you want my detailed protocol, send me a PM
it is referring to the w/v percentage of monomer in the total volume of gelling solution. different final volumes do not make a difference as long as all volumes are modified proportionately.
thanks! very enlightening!
i though that for APS and TEMED proportions should be respectively 1/100 and 1/1000 of the gel volume (ie for 10ml of gel to pour, i add 100µl of APS and 10µl TEMED...)
Try this link