Today, I completed my 1st coIP experiment successfully. However, I have a few questions:
I had two proteins of interest (A & B) and I wanted to see if Protein C pulls it down. I found that Protein C pulls down protein B (25 KDa), but not protein A (~70 kDa).
What could be the case for not detecting A? Is it the antibody or there is no/weak interaction. A recent interactome study has shown that A & C interact and have a good interaction.
I saw bands in my 2 inputs and 2 samples but I couldn't see any bands in my IgG. Does this mean my coIP was ok? What if bands were detected in the IgG controls?
Lastly, once I prepared the beads, I added 25 µl of Sample buffer and I loaded the same to my gels. Is there any way to store the beads with or without sample buffer for longer time so that it can be re-used in case my experiment fails?