NaCl concentration and chromatin - (Aug/11/2005 )
For a native ChIP (no crosslinking) I prepared oligo-nucleosomes (mostly mononuceosomes) with MNAse. The NaCl conc. of the samples is about 100mM. Before the incubation with antibodies, the protocol directs me to dilute to 20mM NaCl. How can I avoid this dilution since DNA concentration is relatively low? What is the molecular background of this 20mM NaCl?
Thanks for every reply!
I think increased NaCl concentration means higher stringency. As you see most of your wash buffers have the higher salt concentration as it interferes with the hydrophobic interactions between antigen and antibody. As It has to be emperically determined for different antibodies it might work for you at 100mM NaCl. As a safe measure generally protocols give sub minimal concentrations of NaCl for pulldown and increase the concentration during washes.