Problem:
I've just been introduced to the procedure of Western blotting from my reading, though I'm not entirely sure about some points. I'd appreciate it if someone could help me with this.
Required:
Question 1: What exactly is the function of the blocking buffer?
Question 2: Why does one need a secondary antibody?
Question 3: When discussing nitrocellulose membranes, say of a 0.45 5m pore size, what does 5m refer to?
Could someone explain this to me?