Question about binding studies

Shyam Gunti wrote:


What cells and type of oil are you using? I have made good experience
with butyl phtalate for erythrocytes.


Prenyl-groups are very hydrophobic and will partition into the lipid
bilayer. Thus non-specific binding is bound to be significant. You could
try to treat the cells with a mixture of proteases to strip them of
receptor. Any remaining binding would be unspecific.


As long as the ligand is not internalised or digested: yes. But if the
binding is quick, why incubate for a long time?


You could label the ligand with an environmentally sensitive fluorescent
probe. Transfer from water to a hydrophobic binding pocket would
increase fluorescence intensity. Binding would also change fluorescence
polarisation due to the lower rotational freedom (if you have access to
the necessary equipment). Both effects can be used to monitor binding in
real-time.