After immobilization, I have tried to equilibrium the system by injecting the running buffer at flow rate of 60ul/min for 40 mins. The sensogram 2-1 (2-protein , 1-blank as reference) shows spikes on every 500s. Isn't strange?, Both Fc-1 and Fc-2 also looks similar. there were spikes in every 500th second. What I can get from this. After this also I could not avoid the spikes in the pump strokes at the time of experiments. I gave 5 mins of stabilization time for every cycle, still the spikes appeared again.
It is not strange. Your pump is 500 ul and flow is 60 ul so every (500/60)*60 seconds the has to be refilled. Then the flow stops for some seconds and because of that (and because ch1 en 2 are in series) the spike appears. You cannot avoid them.
Your system looks properly equilibrated. When you perform your experiment with double referencing (reference subtraction and blank injection subtraction;
) your data will be fine.