Thanks for your comment, Well, I did EDC/NHS activation on channel 1(in which I immobilize the protein) and did ethanol amine capping on both channels. I used, SensiQ gold only chips from icx nomadics. SAM was prepared by myself.
Do I need to do EDC/NHS activation on both channels?
The activation does what it say. Activation of the sensor surface so it can bind other molecules. In case of the NHS/EDC the carboxyl groups of your SAM are modified so that the next injection with primary amines (such as ethanol amine) will bind. Without activation no immobilization.
Thanks for your comments. So far I have been activating with EDC/NHS only in channel one (where I Immobilize the protein) and I did the ethanol amine injection on both channels. I guess as per your comments, I have to activate both channels ( including reference channel ) with EDC/NHS and then de-activate with ethanol amine on both channels right? Let me do in this way and see what happens.
I am trying to use SPR to study the interaction of small molecules ( my compounds are hydrophobic and sticky) with its binding protein. I am getting too much of nonspecific binging on the reference channel, so its hard to extract the specific binding. I used a mixed carboxyl and hydroxyl SAM to bind the protein Does anyone got experience with these kinds of molecule interaction using SPR?