Hi Sudip,

In general, co-IP conditions should mimic the intracellular conditions for different protein complexes in a cell solution. The protein-protein interactions are often transient and weak interactions between short amino acid sequences in different proteins. It is therefore important to optimize the binding conditions in terms of salt and detergent (see ref Alber et al., (2007) Determining the architectures of macromolecular assemblies Nature 450, 683-694).

Components that can be varied to

increase or decrease stringency of the Co-IP include;

• Salts

• Detergents

• Dithiothreitol (DTT)

• Protease Inhibitors

Buffer modifiers

• Sodium Chloride (e.g., 1 M NaCl)

• Dithiothreitol (e.g., 1 M DTT)

• Magnesium Chloride (e.g., 1 M MgCl2)

• Potassium Acetate (e.g., 1 M KOAc)

• TweenR-20

• TritonR X-100

• Protease inhibitors without EDTA

kind regards

ketil