I looked for several books and protocols, however I couldn't figure out.
I have high background in IP and there is not even candidate band either.
I didn't know the result is right because I don't know how to do IP very well.
In the protocols, there are just simple words..
"Add lysis buffer 0.5~1ml and centrifuge 2500g and discard sup. Do same washing more than 4 times.
These are what I would like to know
1. Does it matter to use lysis buffer or lysis buffer with inhibitor during washinhg step?
2. do I have to vortex or shake up and down everytime after adding lysis buffer?
3. If I added 1ml buffer, do I have to take out almost 1ml or a lot less after centrifugation?
4. What is efficient way to take out buffer with not touching bead?
Please, help me ... I am stucking in this IP problem for several weeks.