pooling samples

[NepeanBone]

Dear All,
I have the following question re pooling of proteins:
Let say you need to extract proteins from treated and untreated mice having 5 per group, and you need to extract proteins to perform western blot and to quantify expressed proteins, so what is the best way to do it?
1. Extract protein individually, run separate western blots and analyse them individually then calculating an average and SDs + do t-test?
2. Extract protein individually and then after quantifying them mixing equal amounts and running one western 3 times?
3. Pool the sample material and simply do one extraction followed by western blot repeated 3 times from same protein samples?
4. Pool in smaller group (eg: 2) the sample, quantify and mix + one western blot?

Any suggestions please are greatly appreciated
Thnak you

[relaxin]

The best way is your option #1, if you want quantitative data. Unless certain tissues do not give enough protein to work with, then you have to pool the samples from 5 mice.