Negative response units????

Peptide 1 was immobilized on a CM5 chip. Thereafter Peptide 1 was exposed to peptides a, b and c in running buffer with washing steps between each peptide. The sensorgram of running buffer only was subtracted from the sensorgrams obtained for each peptide before analysis (graph attached). Peptide a caused an increased of approximately 5 response units while peptide b showed a decrease of 3 response units and peptide c resulted in a decrease of approximately 6 response units.

What is the significance of negative response units?
Does it indicate any type of interaction?
Would it be valid to conclude that peptide a interacted with the bound peptide 1?
KP

Am I the only one not being able to see the graph?

1) The big jumps at the beginning are probably caused by the bulk in the peptide concentration. Reference subtraction is not that good that it can compensate.

2) Looking at the A5, 100 - 500 - 1000 µM curves I see that the response is getting lower with higher concentration. This can indicate that the reference subtraction is not compensating well the differences between the channels. Especially at this low responses this is critical. If the buffer of the peptides is different from the flow buffer you could try to make a calibration plot to compensate for this. Ref: J Med Chem. 2000 May 18;43(10):1986-92.Biosensor analysis of the interaction between immobilized human serum albumin and drug compounds for prediction of human serum albumin binding levels. Frostell-Karlsson A, et all.

3) For the analysis of the curves you should look at the Scrubber Program of Biologic Software ( www.biologic.com.au/ ). This will really speed up your evaluation and makes it possible to make a calibration plot.

4) For now I cannot say that there is a positive interaction between your peptides. This is not only because of the negative response in the curves, but I would like to see more clear a concentration dependent response with more concentrations and a affinity curve.

I hope you are helped by this answer.

Regards

Arnoud