Interaction Analysis and Controls

Sample injections are typically performed at flow rates of 5-30 pi min using sample concentrations corresponding to the affinity of the interaction (Ad, affinity dissociation constant). A measurement will immediately give an answer as to whether binding occurs or not. Nevertheless, each response should be verified through a surface and a sample control. A second trace on the same sensor chip, chemically modified in the same manner as the active surface (e.g. activated/ deactivated), usually serves as a control channel. However, an optimized reference surface carries an inactive molecule, which is similar to the active molecule and immobilized at the same amount. Negative samples like non-specific proteins or carbohydrates, or inactive extracts, serve as a control in solution. 

For the analysis of crude samples, like extracts or serum, a 1 10 dilution with running buffer is often sufficient to decrease any background to an acceptable level. Non-specLfically bound material or aggregated proteins can usually be removed from the surface by using detergent or 5-50 mM sodium hydroxide. 

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