Hello, I am trying to work out the IP of virion protein with human IgG.
I tried to pull down various conc of purified virions with human IgG with added background H293T cells in either a PBS buffer or a NP40 buffer using protein A plus beads (Pierces). I use glycine PH2.5 buffer to elute. I got all same Ct values for a real time PCR assay afterwards for 0, 250, 25000 copies of virion although it appears the background Ct is equivalent to 250 copies.
Why my non-viron control also got Ct on real time assay? (I have tried many times so it can't be contamination all the time)
Why I did not see pull down for them as 0 to 25000 copies of virion got the same Ct? I also see high background.
the virion and human IgG have been confirmed in the ELISA. I can't make it work. So hope anyone here can give a hint.