I have a question regarding the isotope dilution method for correction of technical variability during extraction and measurements in LC-MS/MS experiments. I am constructing calibration curves based on (analyte area)/(deuterium labeled IS area) to determine the concentration of a given analyte in a given sample.
I am now not sure whether I should plot responses against initial concentration (that is the nM before adding the ISTD) or should I plot the final concentration take into account the dilution caused by additing the ISTD mix? Does adding of ISTD is considered a dilution step or just a way to get the response ratio in a given concentration.
Could you please also share some papers or references from where I can learn more about it?
Thanks a lot for your time. I really appreciate it.