Chromatography Forum

I've had trouble with a few assays I'm working on with an Agilent 6410 where I get a "reverse quadratic curve." Low and high points of the standard curve are up to 20% high where as the middle points of the curve are up to 20% low. Its the mirror opposite of the traditional quadratic curves I've seen with instances of instrument signal saturation.

This is happening with neat standards that haven't even been extracted.

Has anyone ever seen this, and have any suggestions?

Although I do not know the nature of your analyte, I'll make a guess:

This could be an adsorption effect.
I have seen such issues with certain compounds (peptides) sticking to the glass vials. The adsorption reaches saturation at high standard levels, that's why the low calibration points are more affected and therefore underestimated which leads to this "reverse quadratic" curve.

I my specific case I simply added an excess of a different peptide to saturate the vial surface before adding my analyte. This worked pretty well and is one of the rare examples when adding a matrix improved performance.
Of course, using different vials might also have worked.

I hope this helps!