Hi everyone. In our group we work on mitochondrial ribosomal protein and we have been studying the protein-protein interaction using co-IP. We were suggested that co-IP can be done after chemical crosslinking, so that more strigent washing steps can be applied to reduce unspecific binding. We would also like to study if our target protein bind to 12S or 16S rRNA, by first doing in vivo UV crosslinking, followed by co-IP and RNA extraction. My question is:

1. Will the chemical crosslinking or UV crosslinking cause non-direct binding partners (protein or RNA) of our target protein being pulled down as well (the whole ribosome being crosslinked together and pulled down)?

2. If the chemical or UV crosslinking is useful, should my control be a A) KO cell line of my target protein, or B) cells without crosslinking, or C) cell lysate co-IPed with normal IgG?

Thank you very much.