Solvent effect

Hi,

I just wanted to gain some clarity on solvent effect. I was advised by the professor that I am working with that solvent effect was occuring in my experiment. My injections involves differing concentrations of MeOH which he suggested caused a spike at the beginning of the injection and then sudden drop in signal before increasing again during dissociation. This is clearly visible with all injections. It seems like the presence of MeOH during association changes the buffer composition slightly to in comparison to the dissociation period causing this effect. I am trying to understand this and also find a good reference to explain this effect but struggling. Any ideas?

Thanks,
Caroline

Hi Caroline,

Can you tell us a bit more about the system you use?
Injections of methanol does not seem a molecular interaction for me.
maybe add a figure of what you see?

Arnoud

Hi Arnoud,

I am using Hepes buffer as the main running buffer and a Protein G chip. The antibody is injected followed by a drug diluted into Hepes buffer but its originial storage solvent in methanol so each injection of drug would contain traces of methanol. The decrease I am talking about happens at 300 seconds just before dissociation.

Thanks,
Caroline

Hi Caroline,

The methanol during the analyte injection is causing a solvent/bulk effect in the association phase. As you can see in the sensorgram, the effect causes a jump (+) at the beginning and a jump (-) at the end of the analyte injection. The negative spike at the beginning of the dissociation phase is because the subtracted curves do not exactly overlap.
The reference subtraction cannot compensate the solvent effect due to the differences in volume exclusion between the reference (probably proteine G alone) and active channel (protein G + antibody).
You can try double referencing to compensate for volume exclusion or make a calibration plot with methanol without analyte.
Arnoud