microBloom

I ordered the wrong type of Phenol for my lab, we needed Phenol saturated with Tris and I just ordered just regular liquid phenol, so I have now brought it upon myself to sature the Phenol with Tris. I have a protocol which I will write here, after which I have a question or two. The protocol tells me to add equal volumes of 0.5 M TrisHCl pH8, mix, and take off the aqueous phase. Then, add equal volumes of 0.1 M TrisHCl pH8, mix, and take off aqueous phase. Then, add equal volumes of 0.1 M (or 0.05 M) TrisHCl pH8, mix, take off aqueous phase, and messure the pH to make sure it's around 7.6-8. My questions: 1. Should I do this in a beaker that has a stir bar in it, or do it in a large cylinder, cover it with parafilm and mix by flipping it upside down and right side up a couple times? The first way seems safer (since Phenol is so dangerous) but I was told that it really needs to be mixed vigourously and a stir bar might not do the trick. The cylinder seems more dangerous, but if I put enough parafilm on there, would it really hold the liquid? (My total volume will be 1 L) 2. When taking of the aquous phase, whether in a beaker or cylinder, is it ok if I pour of at least the beginning of it so I don't have to pipette off the whole 500 mL, or will the bottom layer pour out as soon as I tip over the beaker/cylinder. I was told that if it's in a cylinder the phases will have better separation than a beaker... opinion on this? 3. At the end, does it matter if for the last saturation step I add 0.1 M or 0.05 M TrisHCl pH8? Should I maybe measure the pH before this and determine from that whether I need more or less Tris? 4. How do I know how much mixing is enough? 5. Should I let it stand for any significant period of time beyond letting the phases separate? Does anyone have any other tips?

I read that monoclonal antibodies are better used for Western blots vs. polyclonal antibodies are used for immunoprecipitation, is this true? But if you were to use a polyclonal antibody for a Western, wouldn't those antibodies still only have to bind to the epitopes present on the protein of interest? Why would there be more unspecific binding? Also, how would you determine if you should better use a mouse, rabbit, or other animal to get the antibodies from?

I don't know much about Western blotting since I haven't used the technique, so please excuse this basic question, but what are the practical applications of using monoclonal vs. polyclonal antibodies?